Journal of Bioscience and Bioengineering
VOL. 107 No. 5, 475 – 487, 2009
www.elsevier.com/locate/jbiosc
REVIEW
Lantibiotics: Diverse activities and unique modes of action
Sikder M. Asaduzzaman,1 and Kenji Sonomoto1,2,⁎
Laboratory of Microbial Technology, Division of Microbial Science and Technology, Department of Bioscience and Biotechnology, Faculty of Agriculture,
Graduate School, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan 1 and Laboratory of Functional Food Design,
Department of Functional Metabolic Design, Bio-Architecture Center, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan 2
Received 3 September 2008; accepted 9 January 2009
Lantibiotics are one of the most promising alternative candidates for future antibiotics that maintain their antibacterial
efficacy through many mechanisms. Of these mechanisms, some modes of activity have recently been reported, providing
opportunities to show these peptides as potential candidates for forthcoming applications. Many findings providing new insight
into the detailed molecular activities of numerous lantibiotics are constantly being uncovered. The combination of antibiotic
mechanisms in one lantibiotic molecule shows its diverse antimicrobial usefulness as a future generation of antibiotic. Since
lantibiotics do not have any known candidate resistance mechanisms, the discovered distinct modes of activity may
revolutionize the design of anti-infective drugs through the knowledge provided by these super molecules. In this review, we
discuss the rising assortment of lantibiotics, with special emphasis on their structure-function relationships, addressing the
unique activities involved in their individual modes of action.
© 2009, The Society for Biotechnology, Japan. All rights reserved.
[Key words: Lantibiotics; Structural variants; Structure-activities; Modes of action; Lipid II; Drug design]
ANTIBIOTICS AND RESISTANCE TO ANTIBIOTICS
The discovery of penicillin in 1928 by Alexander Fleming was a
historical milestone in human civilization; the subsequent curing of
individuals with otherwise unbearable and sometimes fatal infectious
diseases by antibiotics has been considered as nothing short of a
medical miracle. The identification and production of a wide variety of
antibiotics on a massive scale have revolutionized medical
approaches. Unfortunately, the initial wide-spread use of antibiotics
has generated a strong evolutionary pressure for the emergence of
resistant bacteria. The exclusive reliance on broad-spectrum antibiotics has further intensified the problem by inducing the development of multi-resistant pathogens. One notorious example is that the
vast majority of the clinical isolates from Staphylococcus aureus strains
have been found to be resistant to methicillin (1). The devastating
threats from acquired resistance to antibiotics are compounding from
all regions of antibiotic end-users. Consequently, there is currently no
antibiotic in clinical use to which resistance has not developed. The
World Health Organization has warned that the rapid increase in
resistance among pathogens may become untreatable (WHO/41.
http://www.who.int 2000). Thus, there is a pressing need to discover
and/or develop new agents that are active even against the emerging
resistant bacteria.
The discovery of new classes of antibacterial compounds based on
targets identified from bacterial genomics is historically invaluable as
a source of antibacterial drugs (e.g., glycopeptides) that bacteria use as
⁎ Corresponding author. Fax: +81 92 642 3019.
E-mail address: sonomoto@agr.kyushu-u.ac.jp (K. Sonomoto).
“weapons” against each other. One such glycopeptide antibiotic,
vancomycin, has long been reliable in treating infections caused by
bacteria resistant to several other antibiotics, and is usually reserved
for the treatment of serious infections, including those caused by the
“super bug” methicillin-resistant Staphylococcus aureus (MRSA).
However, even vancomycin-resistant enterococci (VRE) have now
become quite common (2, 3), and this is made more complex by the
spread of vancomycin resistance genes throughout the pathogens.
Therefore, these dramatic increases in antibiotic-resistant pathogens
have stimulated efforts to identify, develop, or design antibiotics that
may be active against multi-resistant pathogen-caused diseases.
DO LANTIBIOTICS SUPERSEDE CONVENTIONAL ANTIBIOTICS?
Some antimicrobials are now being considered as alternative
antibiotics, such as bacteriocins, bacteriophages, probiotics, and
antimicrobial peptides. The attractive features of some of these
molecules, for example, their natural sources, wide range of activities,
ease of production, and the fact that they are not prone to developing
resistance, have interested researchers seeking to develop new
antibiotics. Among these different sources of alternative antibiotics,
lantibiotics appear to be one of the most promising candidates.
Traditional antibiotics usually exert their activities via a specific mode
of action; for example, penicillin interferes with the cross-linking of
two linear polymers by inhibiting the transpeptidase reaction and
aminoglycoside antibiotics (e.g., streptomycin) inhibit protein biosynthesis by combining with the 30S subunit ribosome, whereas
tertracyclines interfere with the binding of aminoacyl-tRNA to the 30S
subunit ribosome and erythromycin prevents the transpeptidation
1389-1723/$ - see front matter © 2009, The Society for Biotechnology, Japan. All rights reserved.
doi:10.1016/j.jbiosc.2009.01.003
476
ASADUZZAMAN AND SONOMOTO
and translocation steps as a result of binding to the 50S subunit
ribosome. Bacteria tend to develop resistance to all classes of these
conventional antibiotics through a relatively simple mechanism. Even
the antimicrobial peptides derived from many organisms, e.g., the
well-studied peptide megainin, are generally based on their single
action of pore formation in the membrane.
In contrast, lantibiotics have quite diverse activities; for
example, nisin and many other structurally related lantibiotics
(e.g., epidermin/gallidermin) use the cell wall precursor lipid II
bound to the membrane as a docking molecule for pore formation
and combine at least two modes of action, i.e., pore formation and
inhibition of cell wall biosynthesis, for antibacterial activity at
nanomolar concentrations (4–6). Hasper et al. (7) recently elucidated the sequestration mechanism resulting from lantibiotic
action, which helps to explain how some small lantibiotics that
cannot span the bilayer of the bacterial membrane can still
maintain a high level of antibacterial activity. Many other
distinctive modes of action are currently known to be unique to
lantibiotics, to which there are no known natural resistance
mechanisms among bacteria. Therefore, we will discuss the
lantibiotics' molecular mechanisms in order to clarify how these
molecules carry out their exceptional activities.
THE LANTIBIOTIC NISIN, THE FOREMOST ANTIBIOTIC WITH
PROMISING FUTURE POTENTIAL
Surprisingly, the history of lantibiotics is older than that of
conventional antibiotics and dates back to a time before the discovery
of penicillin. The first lantibiotic, nisin, was discovered in the 1920s
and has had widespread application as a safe alternative for food
preservation chemical reagents in approximately 50 countries for over
40 years, without natural resistance development (8, 9). Research
regarding lantibiotics has recently gained renewed interest due to the
emergence of clinical isolates that are resistant to antibiotics such as
vancomycin, the last-resort drug that has been used against infections
caused by Gram-positive bacteria for almost 30 years. The N-acyl-DAla-D-Ala moiety of lipid II is involved in the binding of vancomycin,
and vancomycin-resistant bacteria thus remain sensitive to nisin due
to its different binding site (4). Therefore, there has been a rapid and
diverse expansion of research activities towards lantibiotics. Despite
being the oldest known antibacterial agent, the structure of nisin was
not determined until the elegant landmark studies by Gross and
Morell in 1971 (10), and the word “lantibiotic” was just recently coined
in 1988 as an abbreviation for lanthionine-containing antibiotic
peptides (11). Therefore, although the history of lantibiotics is very
old, a new paradigm is emerging due to their potential and enormous
applications to meet the future challenges of developing antibiotics
that can combat emerging pathogens.
J. BIOSCI. BIOENG.,
FEATURES OF LANTIBIOTICS
All organisms have antimicrobial peptides that act as evolutionarily ancient weapons. The diversity of these antimicrobial peptides
is so great that more than 1000 peptides have been included at
http://www.bbcm.univ.trieste.it/∼tossi/antimic.html (described the
antimicrobial peptides). Among these organisms, bacteria are
remarkable producers of antimicrobial peptides. Bacterial-derived
antimicrobial peptides have a large degree of structural and chemical
diversity. Polypeptide antibiotics (e.g., gramicidin and valinomycin)
are synthesized by large, multi-enzyme complexes from building
blocks provided by a variety of cellular processes (12). Recent
advances in bacterial molecular genetics have further contributed to
new insights into peptide antibiotics. Ribosomally synthesized
peptide antibiotics produced by certain bacteria are termed as
bacteriocins (13, 14). Bacteriocins are divided into classes; lantibiotics are class-I bacteriocins that are antimicrobial peptides containing unusual amino acids, such as thioether cross-linked amino acids
in lanthionine and 3-methyllanthionine, and dehydrated amino acids
in 2,3-didehydroalanine (Dha) and (Z)-2,3-didehydrobutyrine (Dhb)
(15, 16). Post-translational modification renders the lantibiotics
biologically active. A large variety of lantibiotic structures, biosynthetic mechanisms, and modes of action have attracted significant
research interest.
Lantibiotics exhibit a number of notable characteristics. They are
ribosomally synthesized, and in most cases the genes involved in
lantibiotic biosynthesis are clustered, designated by the generic locus
symbol lan, with a more specific genotypic designation for each
lantibiotic member (e.g., nis for nisin, nuk for nukacin ISK-1, gdm for
gallidermin). Lantibiotics are found on conjugative transposable
elements (e.g., nisin), on the chromosome of the host (e.g., subtilin)
or on plasmids (e.g., nukacin ISK-1). The gene clusters for the
biosynthesis of representative lantibiotics are depicted in Fig. 1.
Although the gene order, complexity, and transcriptional organization
of the various clusters differ, three genes (lanAMT) have been
identified that are involved in the biosynthesis of all type-A(II) and
type-B lantibiotics, and four genes (lanABCT) are present in all type-A
(I) lantibiotic gene clusters (the grouping of lantibiotics will be
explained below). These essential genes obviously include the
structural genes that encode the precursor peptides for post-translational maturation (prepeptides), which have been designated lanA,
except for subtilin whose structural gene is historically named spaS.
The lanA genes produce prepeptides that have an extension (leader
peptide) of 23–59 amino acids at their N-terminus in addition to the
mature lantibiotic. The sequencing of the lanA genes indicates that the
dehydro amino acids in lantibiotics are the result of the dehydration of
serine and threonine residues to produce dehydroalanine (Dha) and
dehydrobutyrine (Dhb), respectively. Lanthionine and 3-
FIG. 1. Biosynthetic gene clusters of some representative lantibiotics. Genes with similar proposed functions are highlighted in the same pattern. LanB and lanC genes of type-A(I)
lantibiotics are substituted by the gene lanM of type-A(II) lantibiotics. Despite the differences in the gene order, complexity, and transcriptional organization of the clusters, three
genes (lanAMT) are involved in all type-A(II) and type-B lantibiotics, and four genes (lanABCT) are present in all type-A(I) lantibiotic gene clusters for biosynthesis.
VOL. 107, 2009
methyllanthionine rings are then generated by intramolecular conjugate additions of cysteine to these unsaturated amino acids. Though
the exact function of the leader is not yet clear, the suggested possible
functions include export signaling, protection of the producing strain
by keeping the peptides inactive, and providing scaffolds for the posttranslational modification machinery (17, 18).
In type-A(II) lantibiotics, the bifunctional lanM is responsible for
dehydration and the cyclization reactions. In contrast, in type-A(I)
lantibiotics, lanB is involved in the dehydration of Ser and Thr to form
Dha and Dhb, respectively, and lanC codes for the cyclase that
produces lanthionine or 3-methyllanthionine (Fig. 2). The C-terminus
of the lanM enzyme shows 20–27% sequence identity with the lanC
enzyme, but it has no homology with the lanB enzyme. Direct
evidence for the bifunctional role of the lanM enzyme in catalyzing
dehydration and cyclization has been provided by in vitro reconstitution of lctM in lacticin 481 biosynthesis (19). Some lantibiotics also
undergo further post-translational modifications. For example, the
lanD genes encode the enzyme responsible for the formation of AviCys
and AviMeCys; it is likely that the epiD gene in epidermin and mesD in
mersacidin carry out the in vitro decarboxylation of a C-terminal Cys
residue to form AviCys and AviMeCys, respectively (20, 21). Recently,
one of the most post-translationally modified lantibiotics, paenibacillin, has been isolated and identified to show a broader range of
modifications, including N-terminal acetylation (22).
The N-terminal leader peptide is cleaved, and the mature
lantibiotic is then translocated across the membrane. The prepeptide
of type-A(I) lantibiotics is translocated via ATP binding cassette
transporter LanT, and the leader peptide is catalyzed by serine
protease LanP. However, recent reports of the broad substrate
specificity of NisT for nisin biosynthesis have suggested the secretion
of unmodified, partially modified, or fully modified cyclized nisA
prepeptides and non-lantibiotic peptides fused to the leader peptide
of nisA (23). In contrast to the broad specificity of NisT, the processing
enzyme NisP only removes the leader peptide attached to fully
modified nisin. In type-A(II) lantibiotics, LanT has two functions, to
remove the leader peptide and to export the matured peptide. It has
an extra N-terminal cysteine peptidase domain, as compared to LanT
of type-A(I) lantibiotics (24).
Some gene clusters contain a second transport system, which
usually consists of three genes (lanEFG), and is concerned with the
immunity of the producer strains. In addition, another gene, lanI, is
MODES OF ACTION OF LANTIBIOTICS
477
also assumed to be concerned with self-immunity to some lantibiotics
(25). Additionally, two directive genes (lanKR) are often found to be
involved in the regulation of lantibiotic biosynthesis, encompassing an
important two-component sensory system (26).
STRUCTURES AND LANTIBIOTIC GROUPING
Thus far, more than 50 different lantibiotics have been isolated
from Gram-positive bacteria. Lantibiotics are classified by Jung (27) as
types A and B, based on the topology of their structures. Representatives of the lantibiotic structures are presented in Fig. 3. Type-A
lantibiotics are further divided into two subtypes, elongated type-A(I)
and tail and ring region-containing type-A(II), which have different
genetic organizations (28). In type-A(I) lantibiotics, the lanthionine
and 3-methyllanthionine residues are formed by the action of two
distinct enzymes (LanB and LanC), whereas those that are formed by a
single enzyme (LanM) are termed as type-A(II). Type-B lantibiotics,
such as mersacidin, cinnamycin, duramycin, and ancovenin, are more
globular and compact in structure (29).
In addition, a separate subgroup is formed by the twocomponent lantibiotics consisting of two post-translationally modified peptides that individually have little to no activity but
synergistically display strong antibacterial action. At the present,
this emerging subgroup of two-component lantibiotics encompasses
the structurally closely related lacticin 3147, plantaricin W, and
staphylococcin C55, and the completely unrelated streptococcal
cytolysin, which combines bacteriocin and cytolytic activity against
blood cells (30). The molecular mechanisms responsible for the
synergistic effect of two-peptide bacteriocins are not clear at the
present. Generally, the two-peptide lantibiotics work best at
equimolar concentrations (1:1 stoichiometry). However, an alternative classification of lantibiotics has been proposed by comparing
the leader sequences of many lantibiotics, which reveals two
different conserved motifs other than those presented above. In
this organization (determined by genetics rather than activity
profiles or three-dimensional structures), the class I lantibiotics all
have a common “FNLD” motif between positions -20 and -15 and
usually contain a Pro at position -2. The biosynthetic machinery
involved in the post-translational modifications in this class consists
of LanB and LanC. In contrast, class II peptides contain a
characteristic “GG” or “GA” cleavage site (historically termed the
FIG. 2. An example of the post-translational maturation process of the lantibiotic nisin A. Specific serine and threonine residues (bold) in the nisin prepeptides are dehydrated by NisB.
The cyclization of dehydrated amino acids with cysteine residues is catalyzed by NisC in a regio- and stereo-specific manner, and the protease NisP then proteolytically cleaves the
leader peptide to render the lantibiotic active (for a review of the enzymatic processes involved, see ref. 28).
478
ASADUZZAMAN AND SONOMOTO
J. BIOSCI. BIOENG.,
FIG. 3. Structures of a few lantibiotics. A-S-A, lanthionine; Abu-S-A, 3-methyllanthionine; Dha, dehydroalanine; Dhb, dehydrobutyrine; D-A, D-alanine. Based on the topology of their
structures, lantibiotics are classified into three major groups (27), (A) elongated type-A(I); (B) tail and ring region-containing type-A(II); and (C) globular type-B lantibiotics. In
addition, (D) two-component and (E) some irregularly shaped lantibiotics have also been isolated and identified.
“double Gly motif”), contain multiple Asp and Glu residues, and are
usually processed by one modification enzyme (LanM).
ENGINEERING OF LANTIBIOTICS TO DETERMINE THE FUNCTIONS
OF UNUSUAL STRUCTURES
Lanthionine/methyllanthionine bridges are the most notable
features of lantibiotic peptides. These peptides are characterized by
their high contents of unusual amino acid residues that form a
thioether bridge to produce lanthionine and 3-methyllanthionine and
also contain the unsaturated amino acid residues Dha and Dhb (Fig.
3), which are mostly modified forms of serine, threonine, or cysteine
residues. It is now well established, from studies of different
lantibiotics, that these unusual amino acids play a vital role in the
stability and activity of these antibiotic peptides. The mutagenesis of
lantibiotic structural genes has shown the feasibility of changing the
lantibiotic structure by genetic engineering. For example, the removal
of dehydro amino acids, i.e., the replacement of dehydrated amino
acid residues in lantibiotics with other amino acids, reduces their
antibacterial activity. The mutant Dha5Ala has activity against
vegetative cells similar to that of wild-type nisin, but the activity
against spores is nearly abolished (31). The removal of Dha33 by Ala or
the change of Dha5 and Dha33 with Ala leads to a remarkable
decrease in activity, to about 1% of the activity of wild-type nisin (32).
The replacement of Dha by Dhb and vice versa has been reported for
many lantibiotics. The formation of Dhb instead of Dha in the
structural region at position 5 of the nisZ gene led to the production
of mature nisin Z that shows 2–10 fold lower antibacterial activity
(50–90% less than that of the wild-type) against many indicator strains
VOL. 107, 2009
(33, 34). The Dhb10Dha mutant of mutacin II was reported to have
similar activity (35), whereas the mutant Dhb14Dha did not show any
noticeable change in gallidermin activity (34). The replacement of Dhb
by Ala at positions 16 and 20 of Pep5 is found to lower the activity
toward some indicator strains (36). The mutant Dha16Ile in mersacidin causes a great reduction in its activity against M. luteus and S.
pyogenes (37).
As with the other lantibiotics mentioned above, the two-peptide
lantibiotic lacticin 3147 also showed a dramatic reduction or
elimination of antimicrobial activity, due to mutagenesis in the
lanthionine bridges. Cotter et al. (38) reported alanine scanning
results that showed that 12 out of the 14 mutations involved in 6 out of
the 7 lanthionine bridges in lacticin 3147 peptides result in
elimination of bioactivity. They also found that changing five
dehydrated residues resulted in a drop in activity. The introduction
of a new thioether bridge in the lantibiotic Pep5 results in a dramatic
decrease in antimicrobial activity (36). The replacement of amino
acids in all positions is not tolerated by the biosynthetic machinery,
and expression does not occur. For instance, an attempt to generate
the mutant Dhb10Ala mutacin II resulted in no detectable mutacin
production (35), and the change of Ser3, Ser19, or Cys22, which form
the lanthionines, also results in a loss of gallidermin production (39).
MODES OF ACTION OF LANTIBIOTICS
TABLE 1. Salient features of some notable structural derivatives of lantibiotics
Lantibiotic
Nisin A
Nisin Z
Due to the importance of the unusual structures in lantibiotics,
structure-activity relationships have been determined by numerous
studies. Some important structural variants from various derivatives,
which show a change in the activities and/or properties of lantibiotics,
are included in Table 1.
Cotter et al. (38) scanned all 59 amino acids of the two-component
lantibiotic 3147 and found that at least 36 retain some bioactivity and
that some of the amino acids cluster to form variable domains within
the peptides. The glutamate residue in the A-ring of the lacticin 481
subgroup and in the B-ring of mersacidin is conserved and is critically
important for activity, but it has been shown to be nonessential in
lacticin 481 (37, 40).
TARGET SELECTION AND USE OF A DOCKING MOLECULE
Generally, many lantibiotics (e.g., nisin, nukacin ISK-1) bind to
the membrane, leading to subsequent action. Nukacin ISK-1 binds
the anionic membrane by the lysine residues in the tail region,
which plays a vital role in its antibacterial activity (41). In the case
of nisin, membrane permeabilization occurs after target recognition
and formation of a complex with nisin and lipid II (4) for further
action. Hyde et al. (42) reported that the prime target of nisin in
inhibiting peptidoglycan biosynthesis is near the cell division site.
Epidermin shares a recognition motif with nisin and binds to both
lipid I and lipid II (5). The activity of nisin against vancomycinresistant bacteria is a result of the fact that nisin does not make
contact with vancomycin's binding site (L-Lys-D-Ala-D-Ala moiety of
pentapeptide) on lipid II. Instead, nisin binds the pyrophosphate
moiety of lipid II, allowing this lantibiotic to be effective against
vancomycin-resistant bacteria (43). The antibacterial activities of
plantaricin C are similar to that of nisin; it that strongly inhibits in
vitro lipid II synthesis and forms a stable complex with lipid II,
indicating that both nisin and plantaricin C may target the same
structures in lipid II (44). Smith et al. (45) have recently shown that
mutacin 1140 causes membrane disruption in the artificial membrane and reported that, although it incorporates lipid II, it is
arranged in a manner different than that of the nisin A complex.
The two-peptide lantibiotic lacticin 3147 binds specifically with
lipid II in the outer leaflet of the bacterial cytoplasmic membrane.
Lacticin 3147 A1 (LtnA1) forms a lipid II:LtnA1 complex and another
Derivative
Properties
Activity
Ref.
T2S
N20P/M21V/
K22T/K22S
S3T
Dha instead of Dhb
Change in hinge region
Increased
Enhanced
34
100
Residue for A ring
formation
Change of Dha to A
Dhb instead of Dha
Reduction of positive
charge
Additional cysteine
residue
K increased solubility
Improved solubility
Very low
34
No production
2–10 fold lower
Similar
33
33
34
S5A
S5T
K12P
T13C
M17K
N20K
N20E/
N21E
N20V/N20A
M21K
M21K/Dhb /
K22G
N27K/H21K
Gallidermin
STRUCTURE-ACTIVITY RELATIONSHIPS OF STRUCTURAL VARIANTS
479
Nukacin ISK-1
Fragments and
chimeras
V32E
A12L
K1A-K2AK3A
NisA1–12
NisA1–20
NisA1–29
Lact4816–27
Nis1–11Sub12–32
Negative charge in
hinge region
Change in hinge region
Improved solubility
Change in hinge region
Improved solubility
Influenced C-term.
charge
Ability to form pore
disrupted
Reduction of positive
charge from N-terminal
Rings C, D and
E cleaved
Rings D and E cleaved
All lanthionine
rings retain
N-terminal 5
residues removed
Chimeric peptides from
nisin and subtilin
Inactive/not
101
produced
Reduced
34
Active against Gram- 95
negative bacteria
Inactive
102
Very low
102
Active against Gram- 95
negative bacteria
Very low
102
Similar
3–5 fold lower
102
101
Similar
49
32 fold lower
41
Inactive
103
100 fold lower
10 fold lower
103
103
10 fold lower
104
Similar to nisin,
6–8 fold higher
than subtilin
94
component (LtnA2) recognizes the complex, leading to a high affinity
three-component complex for subsequent action (46). An exchange of
the associated mutant peptide LtnA1-Leu21Ala abolished peptide
production (47), and it is noteworthy that a corresponding leucine is
also found in a number of other lantibiotics within the same subgroup
as LtnA1, i.e., mersacidin, actagardine, and plantaricin W. The
surrounding residues of this leucine are highly conserved. In the
case of mersacidin, it is found to be involved in lipid II binding (48),
and the residue may also be related to lipid II recognition for this
peptide.
TWO-PEPTIDE LANTIBIOTICS WORK SYNERGISTICALLY
A number of two-peptide lantibiotics (those that synergistically
function at optimal concentrations) have been identified during the
last decade, of which lacticin 3147, staphylococcin C55, plantaricin W,
Smb, BHT-A, and haloduracin are closely related. Lacticin 3147 (Fig. 3)
is a well-studied two-peptide lantibiotic with exceptional antibiotic
efficacy that is achieved when two killing mechanisms are combined.
It is also effective against multidrug-resistant pathogens such as MRSA
and VRE. However, some reports have indicated that its significant
activities in the nanomolar concentration range are, to some extent,
strain or species specific. Wiedemann et al. (46) reported that lacticin
3147 peptides (LtnA1 and LtnA2) have a very strong synergistic effect
against Lactococcus lactis, but a remarkably weaker effect against Micrococcus flavus. Interestingly, the A1 peptide and mersacidin are
almost equally effective against the lactococcal strain, but their
activities differ by a factor of 30 against Micrococcus. The activity of
lacticin 3147 involves the binding of the LtnA1 peptide to lipid II. Both
activities (pore formation and inhibition of cell wall biosynthesis)
480
ASADUZZAMAN AND SONOMOTO
J. BIOSCI. BIOENG.,
require the presence of two peptides whose intermolecular interactions appear to be stabilized by lipid II (46).
O'Connor et al. (47) determined the closeness of staphylococcin
C55 to lacticin 3147 and reported that 86% (LtnA1 and C55α) and 55%
(LtnA2 and C55β) of the peptides are identical at the amino acid level.
They also reported that the significance of the relatedness between
these two lantibiotics is so remarkable that the hybrid peptide pairs
LtnA1:C55β and C55α:LtnA2 show activities in the single nanomolar
range, reflecting well with the native pairings. The mutagenesis of the
LtnA1 peptide with the equivalent residues in C55α does not produce
the mutant LtnA1-Leu21Ala. This may be due to the positioning of this
residue in a putative lipid II binding loop.
MODES OF ACTION OF LANTIBIOTICS
The activities of lantibiotics are mostly based on different killing
mechanisms that are combined in one molecule. For example, the
prototypic lantibiotic nisin inhibits peptidoglycan synthesis and forms
pores through specific interactions with the cell wall precursor lipid II
(6). As another example, the mutant [A12L] gallidermin has a
diminished pore formation ability but is as potent as wild-type
gallidermin, indicating that pore formation does not contribute to the
killing of bacteria for this mutant gallidermin (49). It is now well
known that the multiple activities of lantibiotics combine differently
for individual target strains. However, the general steps involved in
lantibiotic activities include i) binding to the bacterial membrane,
followed by insertion into membrane, and ii) the use of receptor/
docking molecules to exert structure-based activity. We will focus, in
detail, on these steps of lantibiotic activities, which are responsible for
their potential antibacterial actions.
BINDING OF LANTIBIOTICS TO MEMBRANE AND
INSERTION INTO MEMBRANE
Many studies have shown that membrane binding is the first step
in lantibiotic modes of action. Altering the charge distributions in
nisin, for example, removing positive charges from the N- or Cterminal region of nisin, hampered the initial interactions of the
peptide to the membrane (50). By comparing the native nisin with its
variants, it was also reported that electrostatic attractions encourage
the initial association of nisin with the membrane. Breukink et al. (51)
reported that the highly positive-charged C-terminus of nisin interacts
primarily with the anionic surface of the bacterial cell membrane. The
same study also indicated a very low association with the anionic
lipids by the Val32Glu mutant of nisin Z and assumed that the
introduction of a negative charge into nisin Z would result in
electrostatic repulsion from the negatively charged phospholipids.
The strongly reduced binding affinity of a nisin1–12 fragment to anionic
phospholipids further indicates the importance of the C-terminus of
nisin for binding to the membrane (52). Although the initial binding
to the membrane surface seems to involve the C-terminus of nisin,
studies with a variant of nisin Z, in which a short peptide is fused into
its C-terminus, show that the C-terminus translocates across the
membrane (53). This translocation of the C-terminus is correlated
with pore-forming activity, and both the activities are dependent on
anionic lipids. Once it is electrostatically bound, the peptide adopts a
membrane-spanning orientation in which the C-terminus of at least
part of the molecules forming the pore is located in the lumen of the
vesicle. However, it is now clear that the N-terminal rings of nisin bind
to the disaccharide-pyrophosphate of lipid II, and the positively
charged C-terminus initially interacts with the head-groups of the
lipids in the membrane bilayer. Nukacin ISK-1 (Fig. 3) also has a net
positive charge and binds strongly to the anionic membrane, and its
potential antibacterial activity is crucially dependent on the Nterminus positive charges (Fig. 4) (41). Demel et al. (54) reported
FIG. 4. (A) Binding affinity of a cationic lantibiotic, nukacin ISK-1, to anionic [1,2dimyristoyl-sn-glycero-3-phospho-rac-(1-glycerol) sodium salt] and zwitterionic (1,2dioleyol-sn-glycero-3-phosphocholine) model membranes determined by a surface
plasmon resonance (SPR) biosensor. Nukacin ISK-1 bound to each of the (a) anionic and
(b) zwitterionic model membranes. (B) Dose-response of nukacin ISK-1 toward the
anionic model membrane. Concentrations of nukacin ISK-1 were (a) 20, (b) 15, and (c)
5 μM. RU, resonance unit (41).
that lacticin 481 (with a net charge of zero) has a higher affinity for
the zwitterionic membrane than nisin, which binds to the anionic
membrane.
PORE FORMATION BY LANTIBIOTICS
Nisin and many other cationic type-A(I) lantibiotics have been well
studied in terms of their modes of action involving cytoplasmic and
artificial membranes (4). Numerous studies prior to the late 1990s
focused on the permeabilization of bacterial cell membranes as the
primary mode of action of nisin and other type-A(I) lantibiotics, which
leads to the release of ions and molecules from the bacteria, eventually
resulting in cell death (55). The pores formed by lantibiotics may have
lifetimes of a few to several hundred milliseconds, with diameters of
up to 2 nm (56). The pores of nisin are somewhat anion selective (51)
and the pores of nisin and Pep5 work only in one direction (rectifying)
(57, 58), whereas gallidermin and epidermin form nonrectifying
channels that are more stable (55). The model membrane systems,
such as planar lipid bilayers and liposomes, have a strong influence on
the efficiency of pore formation (59). It has also been shown in
monolayer studies that antimicrobial activity is well correlated with
the nisin-anionic lipid interaction (55, 59).
The two most well-established mechanisms of pore formation are
the barrel-stave and wedge models. In the barrel-stave mechanism,
the cationic lantibiotic monomers bind to the membrane surface
through electrostatic interactions and are assembled into a preaggregate, and the pores are formed at a certain membrane potential,
where the lantibiotic is perpendicular to the membrane (56). In the
VOL. 107, 2009
case of the wedge model, surface-bound lantibiotic molecules bind
parallel to the membrane surface and generate local strain, bending
the membrane in such a way that the lipid molecules, together with
the lantibiotic, form a pore (60). Chikindas et al. (61) proposed a
model for the orientation of lantibiotics in negatively charged
membranes, in which the relatively elongated nisin molecule lies
parallel to the membrane surface with the positively charged sidechains of amino acids pointing out of the lipid bilayer. In contrast with
this model, which demonstrates the most stable orientation, transient
pore formation may result as a molecule passes through the
membrane by conformational change. The well-established model
for pore formation by the lantibiotic nisin has been presented in
Fig. 5A.
Pore formation by nisin is unique, as compared to that of
vancomycin, teicoplanin, and ramoplanin, in that it subsequently
binds with lipid II, using it as a docking molecule to form a pore that is
stable and highly efficient (62). Figure 5B depicts the structure of lipid
II, which portrays the regions involved in the binding of different
antibiotics. Breukink et al. (4) reported that the presence of lipid II in
the membrane increases the pore-forming efficiency of nisin 1000fold as compared to peptides that do not use lipid II. Lipid II-mediated
pore formation by nisin is so dramatic that the presence of only two
lipid II molecules per 105 phospholipid molecules greatly enhances
the release of dyes from vesicles (4, 5).
Nisin forms highly specific pores through its interaction with
lipid II, and the anion selectivity of nisin in model membrane
systems disappears upon the addition of lipid II (6). Lipid II changes
MODES OF ACTION OF LANTIBIOTICS
481
the orientation of nisin from parallel to perpendicular, with respect
to the membrane surface (63), and is recruited into a stable pore
structure (62). The involvements of manifold molecules in the lipid
II-nisin complex are subsequently sufficient to form a defined pore
of uniform structure (62). Therefore, the lipid II-mediated porecomplex is highly stable and unique, as other cationic antimicrobial
peptides form pores in the membrane that are unstable, transient,
and non-uniform in structure (62, 64). The two-component
lantibiotic lacticin 3147 has also been shown to utilize lipid II in a
sequential manner to form a defined pore. However, a few
lantibiotics, e.g., mersacidin, do not form pores. Many novel findings
of the last few years have uncovered the structure-based activities
of lantibiotics, indicating that pore formation is not the major killing
mechanism of lantibiotics.
LIPID-II TARGETING LANTIBIOTIC ACTIVITIES
Bacteria-specific cell wall precursors, e.g., lipid I and lipid II, are
essential for bacterial cell wall biosynthesis. Many antibiotics bind to
these precursors to interfere with peptidoglycan biosynthesis,
preventing the utilization of these molecules by transpeptidase and
transglycosylase enzymes in building the cross-linked network of the
bacterial cell wall. Vancomycin (a peptide antibiotic) is an example of a
compound that kills bacteria by targeting lipid II and has long been
reliable as an essential antibiotic. Vancomycin binds to the D-Ala-DAla moiety of lipid II, and nisin binds to the disaccharides-pyrophosphate region of lipid II, so nisin is even effective against vancomycin-
FIG. 5. Pore formation by the lantibiotic nisin using lipid II as a docking molecule. (A) Nisin binds to the cell wall precursor lipid II, using it as a docking molecule. The N-terminus of
nisin binds lipid II, while the C-terminus is inserted into the bacterial membrane, subsequently forming a pore to release molecules and ions. (B) Chemical structure of lipid II. NMR
analysis of the lipid II-nisin complex reveals that the N-terminal region of nisin (residues 1–12) encages the pyrophosphate moiety of lipid II with a hydrogen bond network (43).
482
ASADUZZAMAN AND SONOMOTO
resistant strains, though both are confined to targeting lipid II (43). It
has recently been demonstrated that lipid II is the prime target of
several other classes of natural products, including lantibiotics. A
growing number of lantibiotics have been shown to interfere with
peptidoglycan biosynthesis by binding to lipid II, which act differently
on lipid II, where different structures of these compounds are used to
explain the sophisticated modes of action directed by the diverse
structures of lantibiotics.
The prototypic type-A(I) lantibiotic nisin is an elongated amphipathic screw-shaped structure in solution, having a net positive
charge. Initially, its bactericidal action was believed to be predominantly involved in the formation of short-lived pores in bacterial cell
membranes (as mentioned before). During the past few years, a
unique mechanism of action has been shown to be exerted by nisin,
which renders it highly potent against many Gram-positive bacteria
at nanomolar concentrations (4, 6). Many ambiguities have been
clarified on the modes of action of lantibiotics following the report of
Breukink et al. (4), suggesting that nisin interacts in a highly specific
manner with lipid II. The dissimilar sensitivities of lipid-II targeting
lantibiotics to different indicator strains may be due to the presence
of different lipid II contents among various microorganisms (e.g., E.
coli, 2 × 103 molecules per cell; Micrococcus lysodeikticus, 105
molecules per cell) (65, 66). Many studies have subsequently
shown that this inhibition is caused by binding to the lipidassociated peptidoglycan precursors lipid I and lipid II, with lipid II
binding having the more predominant effect (for a review see ref. 67,
68). However, structural information on the interaction of lantibiotics
with the cell wall precursor so far has been restricted to lipid II. Nuclear
magnetic resonance (NMR) data reveal that the pyrophosphate moiety
of lipid II interacts with the backbone amides of rings A and B of nisin
via six hydrogen bonds (43). Bonev et al. (69) reported that nisin can
also bind to bactoprenol pyrophosphate; however, the affinity is
considerably lower than that for the complete lipid II molecule. This
indicates that, for high-affinity binding of nisin, additional interactions
must take place, presumably between the N-acetylmuramyl moieties,
whereas the pentapeptide side chain and the isoprenoid moiety are
not involved. The inference of the interaction of lantibiotics with lipid
I stems mainly from the observation that lipid II biosynthesis is
strongly blocked, but the structural analysis of a lantibiotic-lipid I
complex has not yet been reported. The A and B ring system of
nisin, which has been shown to be responsible for binding with
lipid II, in particular the pyrophosphate moiety, is conserved in
nisin, subtilin, epidermin, gallidermin, and plantaricin C. Bonelli et
al. (49) showed that gallidermin/epidermin has a higher affinity to
lipid II than nisin and suggested that the structural element may be
lysine at position 4 (isoleucine in nisin), which may provide an
additional positive charge to enhance binding to the pyrophosphate
moiety.
Mersacidin, actagardin, and cinnamycin are globular type-B
lantibiotics and also bind to lipid II, but have no structural similarity
with nisin and epidermin (Fig. 3). They act by disrupting the
enzyme function of cell wall biosynthesis, by the formation of a
complex with lipid II (48, 70). Specifically, these compounds
prevent the activity of transglycosylases (70). It is important to
note that mersacidin does not form pores upon binding to lipid II;
this is the reason for its moderate MIC values. However, the
compound is very effective in vivo against staphylococcal infections
(71–73), including MRSA and vancomycin-resistant enterococci
(70). In vitro peptidoglycan synthesis assays suggested that
epidermin and nisin accumulate lipid I, indicating that they may
also inhibit the conversion of lipid I to lipid II (5). The two-peptide
lantibiotic laciticn 3147 works at nanomolar concentrations with a
1:1 stoichiometry (LtnA1:LtnA2). The LtnA1 peptide interacts
specifically with lipid II, which recruits LtnA2 for the inhibition of
cell wall biosynthesis and pore formation (46).
J. BIOSCI. BIOENG.,
CHANGES IN BACTERIAL MORPHOLOGY BY LANTIBIOTICS
The peptidoglycan of bacteria is a dynamic system, which is the
prime target of many lantibiotics, including nisin. Hyde et al. (42)
showed the effects of nisin on B. subtilis cells, which causes rapid
membrane permeabilization and subsequent changes in length, crosssection, shape, and population distributions (Figs. 6 and 7). They
concluded that the lethal action of nisin is due to the concerted effects
of membrane permeabilization, followed by cell wall inhibition and
metabolic deregulation of bacterial division. The principal site of
action for nisin is located in the region of rapid cell wall growth near
the site of septal formation, where the most severe cell wall
malformation occurs (42). Hasper et al. (7) illustrated an action for
lantibiotics by means of a pyrophosphate-mediated mechanism,
through the sequestration of lipid II from sites of bacterial cell wall
synthesis. These findings are consistent in their explanations involving significant aberrations in cell wall morphogenesis, where
bacterial elongation is rapid. B. subtilis cells exposed to nisin form
high numbers of double septa near one another and produce a number
of multiseptal bacteria.
We found that the external morphological appearances of B.
subtilis cells that have been exposed to nukacin ISK-1 are unaltered,
whereas mersacidin-treated cells showed some changes in the overall
morphology. However, cells exposed to nisin show a very different
reaction, which led to a drastic reduction in cell size and abnormal
morphological appearances (Asaduzzaman et al., unpublished data).
The comparison of these lantibiotic-treated ultra-structures showed
that the cells demonstrated large variations in their internal
structures, while showing no change in the inner-structure by nukacin
ISK-1. However, a clear difference was observed in the cross-sections
of nukacin-ISK-1 treated B. subtilis cells, which showed a striking
reduction in cell wall width after addition of nukacin ISK-1 (Asaduzzaman et al., unpublished observation). The most widely studied type-B
lantibiotic, mersacidin, has been reported to cause internal changes in
bacterial cells, resulting in the spreading of chromosomes in the
cytoplasm and ultimately leading to cell lysis (74). In contrast, the
well-known type-A(I) lantibiotic nisin is a lytic-bactericidal agent that
causes multiple aberrations, including leaking of cytoplasmic contents, reduction of cell width, acceleration of cell division, minicell
formation, abnormal morphogenesis of bacterial cells, and eventual
cell death (7, 62).
DISTINCT MODES OF LANTIBIOTIC ACTIONS
We have already described much of the details of different modes of
the lantibiotic actions that are combined in one molecule. For example,
FIG. 6. Transmission electron microscopy observations of bacterial cross-section
projections have been elucidated by Hyde et al. (42). (A) Untreated Bacillus subtilis
cells, in which the cytoplasmic osmotic pressure strengthens the adhesion of the
plasma membrane to the peptidoglycan layer, resulting in circular cross-sections; and
(B) cell wall detachment (indicated by the arrow) from the plasma membrane is visible
after nisin exposure, which relieves the osmotic stress by pore formation, leading to an
astral cross-section after contraction of the plasma membrane. Scale bar: 100 nm.
VOL. 107, 2009
MODES OF ACTION OF LANTIBIOTICS
483
FIG. 7. Hyde et al. (42) observed the morphogenesis of Bacillus subtilis cells. (A1 and A2) Normal progression of septal formation in untreated cells; (B–E) some evidence supporting the
suggestion that the bacterial morphogenesis caused by nisin is a result of morphological aberrations during septation: (B) multiseptal divisions, (C) “corkscrew” cell wall
morphologies, (D) disjointed helical septa, and (E) one example of a division “dead end”, which reduces the bacteria to producing many nonviable “minicells”. Scale bar: 200 nm.
the modes of activity of the prototypic lantibiotic nisin have been
shown to be so sophisticated that its effectiveness as an antibiotic is
gradually increasing upon exploration of its structure-based functions.
Early findings on nisin were mainly confined to the observable
phenomena of pore formation to release molecules and ions (60,
75). Up until the last decade, the advances in the molecular
mechanisms of lantibiotic actions had been very poor. Breukink et al.
(4) were the first to report that peptidoglycan biosynthesis is inhibited
by nisin, and this led to new insights into the molecular mechanisms of
lantibiotic modes of actions. In a later study, Hsu et al. (43) showed that
the nisin-lipid II complex reveals a novel lipid II-binding motif where
the N-terminal backbone amides of nisin coordinate the pyrophosphate moiety of lipid II. Furthermore, the sequestration mechanism
evident from nisin provided insight into how short peptides (e.g.,
gallidermin, epidermin) that may not be capable of spanning the
membrane exert their high antibacterial efficacy. Nisin segregates lipid
II into nonphysiological domains in its mode of action (Figs. 8A and B)
(7). On the other hand, the glycopeptide antibiotic vancomycin does
not segregate lipid II from the cell and clearly produces pools of lipid II
in the septum (Fig. 8C). In agreement with the above findings, Hyde et
al. (42) demonstrated that, in the presence of nisin, septal formation
continues but the bacterial cell displays multiple aberrations, and the
FIG. 8. Hasper et al. (7) illustrated an alternative mechanism of nisin's bactericidal
action, which describes the in vivo segregation of lipid II into nonphysiological domains.
(A) Bacillus megaterium cells incubated with 0.5 μg/ml fluorescein-labeled nisin. The
arrow indicates that the bacterium has already divided. (B) B. subtilis cells incubated
with 4 μg/ml fluorescein-labeled nisin. Fluorescence from nisin appears to be clustered
in patches on the membrane. (C) B. megaterium cells after incubation with 2 μg/ml
labeled vancomycin. The arrows indicate the newly formed division sites or older
exemplars. (D) B. subtilis cells stained with 4 μg/ml fluorescent vancomycin. The
labeled vancomycin reveals pools of lipid II in the septum and as well as lipid II in helical
threads. The insets are Nomarski images.
cell envelope formation is deregulated, leading to aberrant cell
morphogenesis. They also proposed that this mechanism is distinctly
different from the cell wall inhibitory activity of glycopeptides and βlactam antibiotics and also from the actions of pore-forming peptide
antibiotics. In addition to nisin, many other lantibiotics (e.g.,
gallidermin, subtillin, mersacidin) use lipid II but have distinctive
structure-based activities (5, 49, 70). A notable example of the
molecular modes of action has been elucidated for two-peptide
lantibiotics, e.g., laciticin 3147, the well-studied two-component
lantibiotic that works in a sequential manner, where the LtnA1 peptide
interacts specifically with lipid II, then the LtnA2 peptide recognizes
the LtnA1-lipid II complex for pore formation and peptidoglycan
biosynthesis inhibition (46, 76).
STRUCTURAL VARIANTS TO STUDY MODES OF ACTION
The mutants and fragments generated by site-directed mutagenesis and chemical and enzymatic digestion from many works have
provided enormous information regarding the modes of action of
lantibiotics. The introduction of an additional positive charge in nisin
by the Val32Lys variant has a relatively small effect, whereas a
negative charge (Val32Glu) results in about a 4-fold decrease in
activity against some indicator strains (6). Epilancin K7 shares a very
similar C-terminus double-ring system with nisin, which does not
show interaction with lipid II (5). This evidence supports the relatively
unimportant role of the C-terminus of these lantibiotics in biological
activity. However, many studies have strongly suggested that the Nterminus of nisin is essential for binding. For example, a nisin1–12
fragment has no bactericidal activity but shows antagonistic activity
against nisin's bactericidal activity (31), indicating that the fragment
competes with nisin for the binding site. Complete proteolytic
deletion of the D and E rings of nisin leads to a 100-fold decrease
(99% eliminated) in activity (31), whereas chemical disruption of
Dha5, which opens the A ring, results in more than a 500-fold
reduction (less than 0.2%) in antibacterial activity as compared to its
native form (77).
The lipid II variant containing a shorter prenyl tail (3 from 11
isoprene units) can form a complex with nisin, and the length of this
isoprene tail does not affect its pore-forming activity (62). Intermolecular hydrogen bonds between the amides of Dhb2, Ala3, Ile4,
Dha5, and Abu8 on nisin and the oxygens of the pyrophosphate group
of lipid II maintain the pyrophosphate moiety of lipid II within the
cavity. Additionally, MurNAc (N-acetylmuramic acid, a component of
glycan chains) and the first isoprene unit form the binding site for the
recognition of nisin (Fig. 5B). The replacement of Lan in the A ring of
nisin with MeLan resulted in 50-fold reduced affinity of the peptide to
lipid II (6), and it is now well established that chemical opening of the
484
ASADUZZAMAN AND SONOMOTO
A ring causes a nearly complete loss of activity (77). Further
information regarding the binding of nisin and epidermin to both
lipid I and lipid II have been revealed by their NMR structure, in which
both the peptides share a recognition motif (5). Glutamate is
conserved in the A ring of the lacticin 481 subgroup and the B ring
of mersacidin, but it is not required for the activity of lacticin 481 (40).
This discovery indicates that, since this residue is critically important
for the A ring of mersacidin (37), lacticin 481 may have a different
target or may recognize lipid II in a different manner.
INHIBITION OF SPORE GERMINATION
Most studies have mainly focused on the antibacterial activities
against vegetative cells. Nisin, subtilin, and sublancin inhibit the
spores' outgrowths from Bacillus and Clostridium species (78, 79). It
has been proposed that this activity is a result of covalent modification
of a target on the spore coat by nucleophilic attack on Dha5, in the case
of nisin and subtilin (80). The reactive thiol groups on the exterior of
the spores from Bacillus cereus react with compounds such as Snitrosothiols and iodoactetate, and nisin interferes with the modification of these sulfhydryl groups (81), suggesting that the target of nisin
for the inhibition of spore germination is provided by these reactive
thiol groups (82). However, a covalent mechanism has not yet been
established. The replacement of Dha5 by Ala via site-directed
mutagenesis of both subtilin (80, 83) and nisin (31) abolished the
inhibition of spore germination, which indicates it as their putative
site of attack. The above studies clearly suggest that the inhibition of
spore germination is a different lantibiotic activity. Therefore, the
inhibition of spore outgrowth is another distinct biological activity of
lantibiotics, with a different structure-function relationship.
FURTHER BIOLOGICAL FUNCTIONS
Many lantibiotics have interesting biological activities in addition
to their antibacterial activity. The SapB peptide (Fig. 3) produced by
Streptomyces coelicolor works as a morphogenic peptide, and the
novel lantibiotic sublancin (Fig. 3) exhibits lipid II-independent modes
of action, such as the induction of autolysis of staphylococci (79).
Cinnamycin (Fig. 3) and duramycin strongly inhibit the phospholipase
A2 by sequestering its phosphatidylethanolamine (PE) (for multiple
activities, see review 84), in addition to their bactericidal and
hemolytic activities (85, 86). Cinnamycin induces transbilayer lipid
movement, seemingly in a PE-dependent fashion (87).
Nisin and Pep5 also induce autolysis of certain staphylococcal
strains, primarily by breaking down the cell wall at the septa of the
dividing cells, in addition to their usual modes of action (88, 89). The
positively charged lantibiotics associate with the negatively charged
teichoic and lipoteichoic acids, which displace and activate N-acetylL-alanine amidase and N-acetylglucosaminidase enzymes (88, 89).
Though most lantibiotics are reported to use lipid I or lipid II as
their docking molecule, not all lantibiotics bind to these, as
discussed earlier. In most cases, the molecules or mechanisms
involved in the activity have not yet been identified. Pep5 and
epilancin K7 have specifically been shown to not bind lipid I or lipid
II (90), but these lantibiotics still show activities against some
bacteria that are far greater than those of other pore-forming
lantibiotics. The high activity of Pep5 at nanomolar concentrations
against Staphylococcus simulans and S. carnosus signifies that it
employs a different high-affinity receptor or docking molecule for
its potent biological activity (5, 89).
In contrast with lantibiotics, conventional antibiotics do not have
multiple functions in one molecule and do not possess such unique
mechanisms of action. Until now, the molecular structure-based
functions of only a few lantibiotics have been well clarified. Lantibiotic
research is now in an advanced stage, and it is expected that more
J. BIOSCI. BIOENG.,
unique modes of action will be revealed, with a new era of amazing
structure-based antibacterial activities.
RATIONAL AND DE NOVO DESIGN OF LANTIBIOTICS TO
REVOLUTIONIZE ANTIBIOTIC REPERTOIRES
The discoveries of the mechanisms involved as individual lantibiotics work as a novel antibacterial, for example, the recent
discoveries of lipid II as a target for nisin and, in particular, the studies
of the pivotal role played by the pyrophosphate group, have brought
nisin into the forefront as a candidate capable of combating resistant
human infections, as a model case for the design of new antibiotics.
Furthermore, the insights regarding the segregation of lipid II into
non-physiological domains (7) elucidate how small lantibiotic peptides act strongly in vivo by a sequestration mechanism. While it was
previously speculated that NisBTC enzymes had limited specificity, it is
now clear that NisT and NisB have a broad substrate specificity. The
independent functions of NisB, NisC, NisT, and NisP (for a review, ref.
91) present possibilities for designing new lantibiotics. The design of
lantibiotics with respect to modification and export may be possible,
based on the findings that NisB-modified peptides can be produced via
the Sec or Tat system and then cyclized by the in vitro action of NisC
(92). The use of lantibiotic synthetases offers much potential for
designing new peptides. For example, Levengood et al. (93) have
recently demonstrated the use of LctM in making thioether-containing
analogs of enkephalin, contryphan, and inhibitors of human tripeptidyl peptidase II and spider venom epimerase. The versatile catalyzing
capacity of lantibiotic synthetases can thus provide an approach to
prepare libraries of peptides containing thioether rings and/or
dehydro amino acids to overcome the inefficiency of synthetic
chemistry. In addition, the design of modified peptides combined
with different lantibiotics has also been explored (94). Furthermore,
the enzymatic actions of lantibiotics' immunity, processing, and
transportation in combination with its structure-based modes of
action [for example, (i) the presence of lysines in the hinge region of
nisin, which increases nisin's activity in killing Gram-negative bacteria
(95), and (ii) the importance of N-terminal lysines in nukacin ISK-1 for
its membrane binding and activity (41)] may aid in the design of
potential lantibiotics in the future. Moreover, in vitro reconstitutions of
lantibiotics are also in progress in order to revolutionize lantibotics for
enormous applications in the near future.
APPLICATIONS AND FUTURE OUTLOOK
The fact that nisin has no known toxicity to humans has placed it in
a unique position of world-wide acceptance as a powerful and safe
food additive in the control of food spoilage, with widespread
application as a food preservative in almost 50 countries for over
40 years. Nisin has been added to the positive list of food additives by
the European Union (EU) and has also been approved by the Food and
Drug Administration (FDA) (8, 9). Though the proteolytic breakdown
of nisin in the gastrointestinal tract and its low stability at
physiological pH levels limits the initial applications of nisin, nisin
and many other lantibiotics are now being used in agricultural,
veterinary, and, more recently, personal care products. Nisin, mutacin,
mersacidin, etc., are in the preclinical stages of medical application
(96). However, the most significant application of lantibiotics may be
in the treatment of antibiotic-resistant pathogens. Ryan et al. (97)
have reviewed the potential biomedical applications of lantibiotics in
clinical and veterinary therapies. Some notable points are: nisin is
effective against bacterial mastitis, oral decay and enterococcal
infections and is effective in peptic ulcer treatment, treatment of
enterocolitis, etc.; mersacidin and actagardine show remarkable
activity against Staphylococcus aureus including MRSA, bacterial
mastitis, oral decay, acne, etc.; gallidermin and epidermin are effective
VOL. 107, 2009
MODES OF ACTION OF LANTIBIOTICS
against acne, eczema, follicultis, and impetigo and can also be used for
personal care products; mutacin 1140 may prevent dental cavities;
lacticin 3147 is reported to prevent bacterial mastitis, MRSA and
enterococcal infections, prevents oral hygiene, and acne; cinnamycin
may be used for inflamation, viral infection treatment, and blood
pressure regulation; and duramycin and ancovenin can be used for
inflamation and blood pressure regulation, respectively. Some more
remarkable applications of nisin have also been reported, which
include the inhibition of experimental vascular graft infection caused
by methicillin-resistant Staphylococcus epidermidis (98), and more
interestingly, nisin inhibits sperm motility, showing its potential as a
contraceptive agent (99). As there has been a recent threat of the use
of spores of Bacillus anthracis in bioterrorism, the inhibitory activities
of lantibiotics, such as subtilin (80, 83) and nisin (31), against spore
germination may have interesting and potential future applications.
In the post-genomic era, the combined knowledge of genetics,
chemistry, and other approaches can promote new innovations in
lantibiotics. The systematic research of lantibiotics may further
resolve the existing difficulties and demonstrate potential use in
food agriculture as well as in medical fields.
ACKNOWLEDGMENTS
Our work is partly supported by grants from “The Japan Society for
the Promotion of Science (JSPS)”.
References
1. Roder, B. L., Wandall, D. A., Frimodt-Moller, N., Espersen, F., Skinhoj, P., and
Rosdahl, V. T.: Clinical features of Staphylococcus aureus endocarditis: a 10-year
experience in Denmark, Arch. Inter. Med., 159, 462–469 (1999).
2. Weigel, L. M., Clewell, D. B., Gill, S. R., Clark, N. C., McDougal, L. K., Flannagan, S.
E., Kolonay, J. F., Shetty, J., Killgore, G. E., and Tenover, F. C.: Genetic analysis of a
high-level vancomycin-resistant isolate of Staphylococcus aureus, Science, 302,
1569–1571 (2003).
3. Novak, R., Henriques, B., Charpentier, E., Normark, S., and Tuomanen, E.:
Emergence of vancomycin tolerance in Streptococcus pneumoniae, Nature, 399,
590–591 (1999).
4. Breukink, E., Wiedemann, I., van Kraaij, C., Kuipers, O. P., Sahl, H. G., and de
Kruijff, B.: Use of the cell wall precursor lipid II by a pore-forming peptide
antibiotic, Science, 286, 2361–2364 (1999).
5. Brötz, H., Josten, M., Wiedemann, I., Schneider, U., Götz, F., Bierbaum, G., and
Sahl, H. G.: Role of lipid-bound peptidoglycan precursors in the formation of pores
by nisin, epidermin and other lantibiotics, Mol. Microbiol., 30, 317–327 (1998).
6. Wiedemann, I., Breukink, E., van Kraaij, C., Kuipers, O. P., Bierbaum, G., de
Kruijff, B., and Sahl, H. G.: Specific binding of nisin to the peptidoglycan precursor
lipid II combines pore formation and inhibition of cell wall biosynthesis for potent
antibiotic activity, J. Biol. Chem., 276, 1772–1779 (2001).
7. Hasper, H. E., Kramer, N. E., Smith, J. L., Hillman, J. D., Zachariah, C.,
Kuipers, O. P., de Kruijff, B., and Breukink, E.: An alternative bactericidal
mechanism of action for lantibiotic peptides that target lipid II, Science, 313,
1636–1637 (2006).
8. Delves-Broughton, J.: Nisin and its uses as a food preservative, Food Technol., 44,
100–117 (1990).
9. Delves-Broughton, J., Blackburn, P., Evans, R. J., and Hugenholtz, J.: Applications of the bacteriocin, nisin, Antonie van Leeuwenhoek, 69, 193–202 (1990).
10. Gross, E. and Morell, J. L.: The structure of nisin, J. Am. Chem. Soc., 93, 4634–4635
(1971).
11. Schnell, N., Entian, K. D., Schneider, U., Götz, F., Zahner, H., Kellner, R., and Jung,
G.: Prepeptide sequence of epidermin, a ribosomally synthesized antibiotic with
four sulphide-rings, Nature, 333, 276–278 (1988).
12. Mootz, H. D. and Marahiel, M. A.: Biosynthetic systems for nonribosomal peptide
antibiotic assembly, Curr. Opin. Chem. Biol., 1, 543–551 (1997).
13. Nissen-Meyer, J. and Nes, I. F.: Ribosomally synthesized antimicrobial peptides:
their function, biogenesis, and mechanism of action, Arch. Microbiol., 167, 67–77
(1997).
14. Tagg, J. R., Dajani, A. S., and Wannamaker, L. W.: Bacteriocins of gram-positive
bacteria, Bacteriol. Rev., 40, 722–756 (1976).
15. de Vos, W. M., Kuipers, O. P., van der Meer, J. R., and Siezen, R. J.: Maturation
pathway of nisin and other lantibiotics: post-translationally modified antimicrobial peptides exported by gram-positive bacteria, Mol. Microbiol., 17, 427–437
(1995).
485
16. Sahl, H. G., Jack, R. W., and Bierbaum, G.: Biosynthesis and biological activities of
lantibiotics with unique post-translational modifications, Eur. J. Biochem., 230,
827–853 (1995).
17. Jung, G.: Lantibiotics — ribosomally synthesized biologically active polypeptides
containing sulfide bridges and α,β-didehydroamino acids, Ang. Chem., Intl. Ed.
Engl., 30, 1051–1068 (1991).
18. van der Meer, J. R., Rollema, H. S., Siezen, R. J., Beerthuyzen, M. M., Kuipers, O.
P., and de Vos, W. M.: Influence of amino acid substitutions in the nisin leader
peptide on biosynthesis and secretion of nisin by Lactococcus lactis, J. Biol. Chem.,
269, 3555–3562 (1994).
19. Xie, L., Miller, L. M., Chatterjee, C., Averin, O., Kelleher, N. L., and van der Donk,
W. A.: Lacticin 481: In vitro reconstitution of lantibiotic synthetase activity,
Science, 303, 679–681 (2004).
20. Majer, F., Schmid, D. G., Altena, K., Bierbaum, G., and Kupke, T.: The flavoprotein
MrsD catalyzes the oxidative decarboxylation reaction involved in formation of the
peptidoglycan biosynthesis inhibitor mersacidin, J. Bacteriol.,184,1234–1243 (2002).
21. Schmid, D. G., Majer, F., Kupke, T., and Jung, G.: Electrospray ionization fourier
transform ion cyclotron resonance mass spectrometry to reveal the substrate
specificity of the peptidyl-cysteine decarboxylase EpiD, Rapid. Commun. Mass
Spectrom., 16, 1779–1784 (2002).
22. He, Z., Yuan, C., Zhang, L., and Yousef, A. E.: N-terminal acetylation in
paenibacillin, a novel lantibiotic, FEBS Lett., 582, 2787–2792 (2008).
23. Kuipers, A., de Boef, E., Rink, R., Fekken, S., Kluskens, L. D., Driessen, A. J. M.,
Leenhouts, K., Kuipers, O. P., and Moll, G. N.: NisT, the transporter of the
lantibiotic nisin, can transport fully modified, dehydrated and unmodified
prenisin and fusions of the leader peptide with non-lantibiotic peptides, J. Biol.
Chem., 279, 22176–22182 (2004).
24. Havarstein, L. S., Diep, D. B., and Nes, I. F.: A family of bacteriocin ABC
transporters carry out proteolytic processing of their substrates concomitant with
export, Mol. Microbiol., 16, 229–240 (1995).
25. Heidrich, C., Pag, U., Josten, M., Metzger, J., Jack, R. W., Bierbaum, G., Jung, G.,
and Sahl, H. G.: Isolation, characterization, and heterologous expression of the
novel lantibiotic epicidin 280 and analysis of its biosynthetic gene cluster, Appl.
Environ. Microbiol., 64, 3140–3146 (1998).
26. de Ruyter, P. G., Kuipers, O. P., Beerthuyzen, M. M., van Alen-Boerrigter, I., and
de Vos, W. M.: Functional analysis of promoters in the nisin gene cluster of Lactococcus lactis, J. Bacteriol., 178, 3434–3439 (1996).
27. Jung, G.: Lantibiotics: a survey, p. 1–34, in: G. Jung, H. -G. Sahl (Eds.), Nisin and
novel lantibiotics, ESCOM, Leiden, The Netherlands, 1991.
28. Nagao, J., Asaduzzaman, S. M., Okuda, K., Aso, Y., Nakayama, J., and Sonomoto,
K.: Lantibiotics: insight and foresight for new paradigm, J. Biosci. Bioeng., 102,
139–149 (2006).
29. Fredenhagen, A., Maerki, F., Fendrich, G., Maerki, W., Gruner, J., Van Oostrum,
J., Raschdorf, F., and Peter, H. H.: Duramycin B and C, two new lanthioninecontaining antibiotics as inhibitors of phospholipase A2, and structural revision of
duramycin and cinamycin, in: G. Jung, H. -G. Sahl (Eds.), Nisin and novel
lantibiotics, ESCOM, Leiden, The Netherlands, 1991, pp. 131–140 (1991).
30. Gilmore, M. S., Segarra, R. A., Booth, M. C., Bogie, C. P., Hall, L. R., and Clewell, D.
B.: Genetic structure of the Enterococcus faecalis plasmid pAD1-encoded cytolytic
toxin system and its relationship to lantibiotic determinants, J. Bacteriol., 176,
7335–7344 (1994).
31. Chan, W. C., Dodd, H. M., Horn, N., Maclean, K., Lian, L. Y., Bycroft, B. W., Gasson,
M. J., and Roberts, G. C.: Structure-activity relationships in the peptide antibiotic
nisin: role of dehydroalanine 5, Appl. Environ. Microbiol., 62, 2966–2969 (1996).
32. Dodd, H. M., Horn, N., and Gasson, M. J.: A cassette vector for protein engineering
the lantibiotic nisin, Gene, 162, 163–164 (1995).
33. Kuipers, O. P., Rollema, H. S., Yap, W. M., Boot, H. J., Siezen, R. J., and de Vos, W.
M.: Engineering dehydrated amino acid residues in the antimicrobial peptide
nisin, J. Biol. Chem., 267, 24340–24346 (1992).
34. Kuipers, O. P., Bierbaum, G., Ottenwälder, B., Dodd, H. M., Horn, N., Metzger, J.,
Kupke, T., Gnau, V., Bongers, R., van den Bogaard, P., Kosters, H., Rollema, H. S.,
de Vos, W. M., Siezen, R. J., Jung, G., Götz, F., Sahl, H. G., and Gasson, M. J.: Protein
engineering of lantibiotics, Antonie van Leeuwenhoek, 69, 161–169 (1996).
35. Chen, P., Novak, J., Kirk, M., Barnes, S., Qi, F., and Caufield, P. W.: Structureactivity study of the lantibiotic mutacin II from Streptococcus mutans T8 by a gene
replacement strategy, Appl. Environ. Microbiol., 64, 2335–2340 (1998).
36. Bierbaum, G., Szekat, C., Josten, M., Heidrich, C., Kempter, C., Jung, G., and Sahl,
H. G.: Engineering of novel thioether bridge and role of modified residues in the
lantibioitc Pep5, Appl. Environ. Microbial., 62, 385–392 (1996).
37. Szekat, C., Jack, R. W., Skutlarek, D., Farber, H., and Bierbaum, G.: Construction
of an expression system for site-directed mutagenesis of the lantibiotic
mersacidin, Appl. Environ. Microbiol., 69, 3777–3783 (2003).
38. Cotter, P. D., Deegan, L. H., Lawton, E. M., Draper, L. A., O'Connor, P. M., Hill, C.,
and Ross, R. P.: Complete alanine scanning of the two-component lantibiotic
lacticin 3147: generating a blueprint for rational drug design, Mol. Microbiol., 62,
735–747 (2006).
39. Ottenwälder, B., Kupke, T., Brecht, S., Gnau, V., Metzger, J., Jung, G., and Göt, F.:
Isolation and characterization of genetically engineered gallidermin and epidermin analogs, Appl. Environ. Microbiol., 61, 3894–3903 (1995).
486
ASADUZZAMAN AND SONOMOTO
40. Patton, G. C., Paul, M., Cooper, L. E., Chatterjee, C., and van der Donk, W. A.: The
importance of the leader sequence for directing lanthionine formation in lacticin
481, Biochemistry, 47, 7342–7351 (2008).
41. Asaduzzaman, S. M., Nagao, J., Aso, Y., Nakayama, J., and Sonomoto, K.: Lysineoriented charges trigger the membrane binding and activity of nukacin ISK-1,
Appl. Environ. Microbiol., 72, 6012–6017 (2006).
42. Hyde, A. J., Parisot, J., McNichol, A., and Bonev, B. B.: Nisin-induced changes in
Bacillus morphology suggest a paradigm of antibiotic action, Proc. Natl. Acad.
Sci. USA, 103, 19896–19901 (2006).
43. Hsu, S. T. D., Breukink, E., Tischenko, E., Lutters, M. A., de Kruijff, B., Kaptein, R.,
Bonvin, A. M., and van Nuland, N. A. J.: The nisin-lipid II complex reveals a
pyrophosphate cage that provides a blueprint for novel antibiotics, Nat. Struct.
Mol. Biol., 11, 963–967 (2004).
44. Wiedemann, I., Bottiger, T., Bonelli, R. R., Schneider, T., Sahl, H. G., and
Martinez, B.: Lipid II-based antimicrobial activity of the lantibiotic plantaricin C,
Appl. Environ. Microbiol., 72, 2809–2814 (2006).
45. Smith, L., Hasper, H., Breukink, E., Novak, J., Cerkasov, J., Hillman, J. D., WilsonStanford, S., and Orugunty, R. S.: Elucidation of the antimicrobial mechanism of
mutacin 1140, Biochemistry, 47, 3308–3314 (2008).
46. Wiedemann, I., Böttiger, T., Bonelli, R. R., Sven, A. W., Hagge, O., Gutsmann, T.,
Seydel, U., Deegan, L., Hill, C., Ross, P., and Sahl, H. G.: The mode of action of the
lantibiotic lacticin 3147 — a complex mechanism involving specific interaction of
two-peptides and the cell wall precursor lipid II, Mol. Microbiol., 61, 285–297
(2006).
47. O'Connor, E. B., Cotter, P. D., O'Connor, P., O'Sullivan, O., Tagg, J. R., Ross, R. P.,
and Hill, C.: Relatedness between the two-component lantibiotics lacticin 3147
and staphylococcin C55 based on structure, genetics and biological activity, BMC
Microbiol., 7, 24 (2007).
48. Hsu, S. T. D., Breukink, E., Bierbaum, G., Sahl, H. G., de Kruijff, B., Kaptein, R.,
van Nuland, N. A. J., and Bonvin, A. M. J. J.: NMR study of mersacidin and lipid II
interaction in dodecylphosphocholine micelles. Conformational changes are a key
to antimicrobial activity, J. Biol. Chem., 278, 13110–13117 (2003).
49. Bonelli, R. R., Schneider, T., Sahl, H. G., and Wiedemann, I.: Insights into in vivo
activities of lantibiotics from gallidermin and epidermin mode-of-action studies,
Antimicrob. Agents Chemother., 50, 1449–1457 (2006).
50. Giffard, C. J., Dodd, H. M., Ladha, N., Horn, S., Machie, A. R., Parr, A., Gasson, M. J.,
and Sanders, D.: Structure-function relations of variant and fragments of nisin
studied with model membrane systems, Biochemistry, 36, 3802–3810 (1997).
51. Breukink, E., van Kraaij, C. R., Demel, A., Siezen, R. J., Kuipers, O. P., and de
Kruijff, B.: The C-terminal region of nisin is responsible for the initial interaction
of nisin with the target membrane, Biochemistry, 36, 6968–6976 (1997).
52. Moll, G. N., Clark, J., Chan, W. C., Bycroft, B. W., Roberts, G. C., Konings, W. N.,
and Driessen, A. J.: Role of transmembrane pH gradient and membrane binding in
nisin pore formation, J. Bacteriol., 179, 135–140 (1997).
53. van Kraaij, C., Breukink, E., Noordermeer, M. A., Demel, R. A., Siezen, R. J.,
Kuipers, O. P., and de Kruijff, B.: Pore formation by nisin involves translocation of
its C-terminal part across the membrane, Biochemistry, 37, 16033–16040 (1998).
54. Demel, R. A., Peelen, T., Siezen, R. J., de Kruijff, B., and Kuipers, O. P.: Nisin Z,
mutant nisin Z and lacticin 481 interactions with anionic lipids correlate with
antimicrobial activity. A monolayer study, Eur. J. Biochem., 235, 267–274 (1996).
55. Benz, R., Jung, G., and Sahl, H. G.: Mechanism of channel-formation by
lantibiotics in black lipid membranes, in: G. Jung, H. -G. Sahl (Eds.), Nisin and
novel lantibiotics, ESCOM, Leiden, The Netherlands, 1991, pp. 359–372 (1991).
56. Sahl, H. G.: Pore formation in bacterial membranes by cationic lantibiotics. In
Nisin and novel lantibiotics, in: G. Jung, H. -G. Sahl (Eds.), ESCOM, Leiden, The
Netherlands, 1991, pp. 347–358 (1991).
57. Kordel, M., Benz, R., and Sahl, H. G.: Mode of action of the staphylococcin like
peptide Pep 5: voltage-dependent depolarization of bacterial and artificial
membranes, J. Bacteriol., 170, 84–88 (1988).
58. Sahl, H. G., Kordel, M., and Benz, R.: Voltage-dependent depolarization of
bacterial membranes and artificial lipid bilayers by the peptide antibiotic nisin,
Arch. Microbiol., 149, 120–124 (1987).
59. Gao, F. H., Abee, T., and Konings, W. N.: Mechanism of action of the peptide
antibiotic nisin in liposomes and cytochrome c oxidase-containing proteoliposomes, Appl. Environ. Microbiol., 57, 2164–2170 (1991).
60. Driessen, A. J., van den Hooven, H. W., Kuiper, W., van de Kamp, M., Sahl, H.
G., Konings, R. N., and Konings, W. N.: Mechanistic studies of lantibioticinduced permeabilization of phospholipid vesicles, Biochemistry, 34, 1606–1614
(1995).
61. Chikindas, M. L., Novak, J., Driessen, A. J., Konings, W. N., Schilling, K. M., and
Caufield, P. W.: Mutacin II, a bactericidal antibiotic from Streptococcus mutans,
Antimicrob. Agents Chemother., 39, 2656–2660 (1995).
62. Breukink, E., van Heusden, H. E., Vollmerhaus, P. J., Swiezewska, E., Brunner, L.,
Walker, S., Heck, A. J., and de Kruijff, B.: Lipid II is an intrinsic component of the
pore induced by nisin in bacterial membranes, J. Biol. Chem., 278, 19898–19903
(2003).
63. van Heusden, H. E., de Kruijff, B., and Breukink, E.: Lipid II induces a
transmembrane orientation of the pore-forming peptide lantibiotic nisin,
Biochemistry, 41, 12171–12178 (2002).
J. BIOSCI. BIOENG.,
64. Ottenwälder, B., Kupke, T., Brecht, S., Gnau, V., Metzger, J., Jung, G., and Göt, F.:
Isolation and characterization of genetically engineered gallidermin and epidermin analogs, Appl. Environ. Microbiol., 61, 3894–3903 (1995).
65. Storm, D. R. and Strominger, J.: Binding of bacitracin to cells and protoplasts of
Micrococcus lysodeikticus, J. Biol. Chem., 249, 1823–1827 (1974).
66. van Heijenoort, Y., Gomez, M., Derrien, M., Ayala, J., and van Heijenoort, J.:
Membrane intermediates in the peptidoglycan metabolism of Escherichia coli:
possible roles of PBP 1b and PBP 3, J. Bacteriol., 174, 3549–3557 (1992).
67. Chatterjee, C., Paul, M., Xie, L., and van der Donk, W. A.: Biosynthesis and mode
of action of lantibiotics, Chem. Rev., 105, 633–684 (2005).
68. Reisinger, P., Seidel, H., Tschesche, H., and Hammes, W. P.: The effect of nisin on
murein synthesis, Arch. Microbiol., 127, 187–193 (1980).
69. Bonev, B. B., Breukink, E., Swiezewska, E., de Kruijff, B., and Watts, A.: Targeting
extracellular pyrophosphates underpins the high selectivity of nisin, FASEB J., 18,
1862–1869 (2004).
70. Brötz, H., Bierbaum, G., Reynolds, P. E., and Sahl, H. G.: The lantibiotic
mersacidin inhibits peptidoglycan biosynthesis at the level of transglycosylation,
Eur. J. Biochem., 246, 193–199 (1997).
71. Kruszewska, D., Sahl, H. G., Bierbaum, G., Pag, U., Hynes, S. O., and Ljungh, A.:
Mersacidin eradicates methicillin-resistant Staphylococcus aureus (MRSA) in a
mouse rhinitis model, J. Antimicrob. Chemother., 54, 648–653 (2004).
72. Chatterjee, S., Lad, S. J., Phansalkar, M. S., Rupp, R. H., Ganguli, B. N., Fehlhaber, H.
W., and Kogler, H.: Mersacidin, a new antibiotic from Bacillus: fermentation, isolation,
purification, and chemical characterization, J. Antibiot. (Tokyo), 45, 832–838 (1992).
73. Limbert, M., Isert, D., Klesel, N., Markus, A., Seibert, G., Chatterjee, S.,
Chatterjee, D. K., Jani, R. H., and Ganguli, B. N.: Chemotherapeutic properties
of mersacidin in vitro and in vivo, in: G. Jung, H. -G. Sahl (Eds.), Nisin and novel
lantibiotics, ESCOM, Leiden, The Netherlands, 1991, pp. 448–456 (1991).
74. Brötz, H., Bierbaum, G., Markus, A., Molitor, E., and Sahl, H. G.: Mode of action of
the lantibiotic mersacidin: inhibition of peptidoglycan biosynthesis via a novel
mechanism? Antimicrob. Agents Chemother., 39, 714–719 (1995).
75. García Garcerá, M. J., Elferink, M. G., Driessen, A. J., and Konings, W. N.: In vitro
pore-forming activity of the lantibiotic nisin. Role of protonmotive force and lipid
composition, Eur. J. Biochem., 212, 417–422 (1993).
76. Morgan, S. M., O'Connor, P. M., Cotter, P. D., Ross, R. P., and Hill, C.: Sequential
actions of the two component peptides of the lantibiotic lacticin 3147 explain its
antimicrobial activity at nanomolar concentrations, Antimicrob. Agents Chemother., 49, 2606–2611 (2005).
77. Chan, W. C., Bycroft, B. W., Lian, L. Y., and Roberts, G. C.: Isolation and
characterisation of two degradation products derived from the peptide antibiotic
nisin, FEBS Lett., 252, 29–36 (1989).
78. Hurst, A.: Nisin, Adv. Appl. Microbiol., 27, 85–123 (1981).
79. Paik, S. H., Chakicherla, A., and Hansen, J. N.: Identification and characterization
of the structural and transporter genes for, and the chemical and biological
properties of, sublancin 168, a novel lantibiotic produced by Bacillus subtilis 168, J.
Biol. Chem., 273, 23134–23142 (1998).
80. Liu, W. and Hansen, J. N.: The antimicrobial effect of a structural variant of
subtilin against outgrowing Bacillus cereus T spores and vegetative cells occurs by
different mechanisms, Appl. Environ. Microbiol., 59, 648–651 (1993).
81. Morris, S. L. and Hansen, J. N.: Inhibition of Bacillus cereus spore outgrowth by
covalent modification of a sulfhydryl group by nitrosothiol and iodoacetate, J.
Bacteriol., 148, 465–471 (1981).
82. Morris, S. L., Walsh, R. C., and Hansen, J. N.: Identification and characterization of
some bacterial membrane sulfhydryl groups which are targets of bacteriostatic
and antibiotic action, J. Biol. Chem., 259, 13590–13594 (1984).
83. Liu, W. and Hansen, J. N.: Enhancement of the chemical and antimicrobial properties
of subtilin by site-directed mutagenesis, J. Biol. Chem., 267, 25078–25085 (1992).
84. Pag, U. and Sahl, H. G.: Multiple activities in lantibiotics—models for the design of
novel antibiotics? Curr. Pharm., 8, 815–833 (2002).
85. Machaidze, G. and Seelig, J.: Specific binding of cinnamycin (Ro 09-0198) to
phosphatidylethanolamine. Comparison between micellar and membrane environments, Biochemistry, 42, 12570–12576 (2003).
86. Märki, F., Hanni, E., Fredenhagen, A., and van Oostrum, J.: Mode of action of the
lanthionine-containing peptide antibiotics duramycin, duramycin B and C, and
cinnamycin as indirect inhibitors of phospholipase A2, J. Biochem. Pharmacol., 42,
2027–2035 (1991).
87. Makino, A., Baba, T., Fujimoto, K., Iwamoto, K., Yano, Y., Terada, N., Ohno, S.,
Sato, S. B., Ohta, A., Umeda, M., Matsuzaki, K., and Kobayashi, T.: Cinnamycin
(Ro 09-0198) promotes cell binding and toxicity by inducing transbilayer lipid
movement, J. Biol. Chem., 278, 3204–3209 (2003).
88. Bierbaum, G. and Sahl, H. G.: Induction of autolysis of staphylococci by the basic
peptide antibiotics Pep 5 and nisin and their influence on the activity of autolytic
enzymes, Arch. Microbiol., 141, 249–254 (1985).
89. Bierbaum, G. and Sahl, H. G.: Autolytic system of Staphylococcus simulans 22:
influence of cationic peptides on activity of N-acetylmuramoyl-L-alanine amidase,
J. Bacteriol., 169, 5452–5458 (1987).
90. Pag, U., Heidrich, C., Bierbaum, G., and Sahl, H. G.: Molecular analysis of
expression of the lantibiotic Pep5 immunity phenotype, Appl. Environ. Microbiol.,
65, 591–598 (1999).
VOL. 107, 2009
91. Lubelski, J., Rink, R., Khusainov, R., Moll, G. N., and Kuipers, O. P.: Biosynthesis,
immunity, regulation, mode of action and engineering of the model lantibiotic
nisin, Cell. Mol. Life Sci., 65, 455–476 (2008).
92. Kuipers, A., Wierenga, J., Rink, R., Kluskens, L. D., Driessen, A. J., Kuipers,
O. P., and Moll, G. N.: Sec-mediated transport of posttranslationally
dehydrated peptides in Lactococcus lactis, Appl. Environ. Microbiol., 72,
7626–7633 (2006).
93. Levengood, M. R. and van der Donk, W. A.: Use of lantibiotic synthetases for the
preparation of bioactive constrained peptides, Bioorg. Med. Chem. Lett., 18,
3025–3028 (2008).
94. Chakicherla, A. and Hansen, J. N.: Role of the lead and structural regions of
prelantibiotic peptides as assessed by expressing nisin-subtilin chimeras in
chemical, and antimicrobial properties, J. Biol. Chem., 270, 23533–23539 (1995).
95. Yuan, J., Zhang, Z. Z., Chen, X. Z., Yang, W., and Huan, L. D.: Site-directed
mutagenesis of the hinge region of nisin Z and properties of nisin Z mutants, Appl.
Microbiol Biotechnol., 64, 806–815 (2004).
96. Breukink, E. and de Kruijff, B.: Lipid II as a target for antibiotics, Nat. Rev. Drug
Discov., 5, 321–332 (2006).
97. Ryan, M. P., Hill, C., and Ross, R. P.: Exploitation of lantibiotic peptides for food
and medical uses, in: C. J. Dutton, M. A. Haxell, H. A. I. McArthur, R. G. Wax (Eds.),
Peptide antibiotics — discovery, mode of action and applications, Marcel Dekker,
New York, 2002, pp. 193–242 (2002).
MODES OF ACTION OF LANTIBIOTICS
487
98. Ghiselli, R., Giacometti, A., Cirioni, O., DellAcqua, G., Mosshegiani, F., Orlando,
F., Damato, G., Rocchi, M., Scalise, G., and Saba, V.: RNA III-inhibiting peptide and/
or nisin inhibit experimental vascular graft infection with methicillin-susceptible
and methicillin-resistant Staphylococcus epidermidis, Eur. J. Endovasc. Surg., 27,
603–607 (2004).
99. Aranha, C., Gupta, S., and Reddy, K. V. R.: Contraceptive efficacy of antimicrobial
peptide nisin: in vitro and in vivo studies, Contraception, 69, 333–338 (2004).
100. Field, D., Connor, P. M., Cotter, P. D., Hill, C., and Ross, R. P.: The generation of
nisin variants with enhanced activity against specific gram-positive pathogens,
Mol. Microbiol., 69, 218–230 (2008).
101. van Kraaij, C., Breukink, E., Rollema, H. S., Bongers, R. S., Kosters, H. A., de
Kruijff, B., and Kuipers, O. P.: Engineering a disulfide bond and free thiols in the
lantibiotic nisin Z, Eur. J. Biochem., 267, 901–909 (2000).
102. Rollema, H. S., Kuipers, O. P., Both, P., de Vos, W. M., and Seizen, R. J.:
Improvement of solubility of the antimicrobial peptide nisin by protein
engineering, Appl. Environ. Microbiol., 61, 2873–2978 (1995).
103. Chan, W. C., Leyland, M., Clark, J., Dodd, H. M., Lian, L. Y., Gasson, M. J., Bycroft,
B. W., and Roberts, G. C.: Structure-activity relationships in the peptide antibiotic
nisin: antibacterial activity of fragments of nisin, FEBS Lett., 390, 129–132 (1996).
104. Uguen, P., Hindre, T., Didelot, S., Marty, C., Haras, D., Pennee, J. P. L., ValleeRehel, K., and Dufour, A.: Maturation by LctT is required for biosynthesis of fulllength lantibiotic lacticin 481, Appl. Environ. Microbiol., 71, 562–565 (2005).