Bacterial DNA restriction and modification systems: structure,
function and evolution.
David Dryden
School of Chemistry
University of Edinburgh
Bacterial DNA restriction and modification systems: structure,
function and evolution.
Current group:
Gareth Roberts, John White, Laurie Cooper,
Ed Bower, Matt Tilling
Collaborators:
University of Cambridge: Robert Henderson, Mike Edwardson
University of Edinburgh: Noreen Murray, Garry Blakely, Malcolm
Walkinshaw, Wilson Poon, Anita Jones
St. George's,
University of London:
Patrick Houston, Jodi Lindsay
University of Leeds:
Chris Kennaway, John Trinick
University of Portsmouth: James Taylor, Geoff Kneale
University of St. Andrews: Jim Naismith
Bacterial DNA restriction and modification systems: structure,
function and evolution.
1.
Restriction and modification (RM) systems and their
inhibition by antirestriction systems.
2.
Type I RM enzymes: structure, function and evolution.
3.
Some speculation on the antiquity of RM systems and their
influence on evolution.
RM systems control Horizontal Gene Transfer.
Tock and Dryden, Curr. Op. Microbiol. (2005) 8, 466.
The major classes of RM systems.
REBASE Genomes
(4910 Bacteria, 191 Archaea)
With RM
Bacteria:
4651
Archaea:
190
System
Type I
Type II
Enzymes
Restriction endonuclease
Separate restriction
and modification
endonuclease and
methyltransferase in one
modification
complex.
methyltransferase.
SAM, ATP, Mg2+ for
Mg2+ for endonuclease.
endonuclease
SAM for methyltransferase.
Cofactors
No RM
259
1
There are 11
subtypes of
Type II systems!
SAM for methyltransferase.
DNA target site
Methylation
Cleavage site
Bipartite, asymmetric.
Various.
AAC (N6) GTGC for EcoKI
GATATC for EcoRV
At recognition site, prefers
At recognition site, no
hemimethylated targets.
preference.
Variable locations from 100
Fixed location at or near to
to 50,000 base pairs from
recognition site.
recognition site.
Also Type III and
Type IV
systems.
The major classes of RM systems are Type I and Type II systems.
The tools of molecular biology are
the simplest Type II restriction
enzymes.
Nature has evolved systems of far
greater complexity such as the
Type I systems.
Type I RM enzymes comprise 5
subunits
1x HsdS binds DNA target
2x HsdM binds SAM, methylates
DNA
2xHsdR hydrolyses ATP,
translocates and cuts DNA
Roberts et al. Nucleic Acids Res. (2003) 31, 1805
Dryden Nature Struct. Mol. Biol. (2005) 11, 804.
+Mg2+
+Mg2+
ATP
SAM
Antirestriction systems such as antirestriction proteins
control RM systems.
They co-evolve to assist Horizontal Gene Transfer.
Ocr from phage T7
Antirestriction methods:
1. Avoid having target sites
2. Modified DNA bases
3. Co-injection of antirestriction proteins
4. Consumption of cofactors for R/M systems
5. Enhance host modification activity
6. Proteolysis of RM enzymes
7. Encode specialised antirestriction proteins
e.g. ArdA and ArdB on conjugative transposons and
plasmids and Ocr on phage T7. Plus many others as
yet poorly characterised.
Bickle and Kruger, Microbiol. Rev. (1993) 57, 434.
Tock and Dryden, Curr. Op. Microbiol. (2005) 8, 466.
Dryden, Trends Biotech. (2006) 24, 378.
Walkinshaw et al. Mol. Cell (2002) 9, 187.
McMahon et al., (2009) Nucl. Acids Res. 37, 4887.
Serfiotis-Mitsa et al., (2009) Nucl. Acids Res. 38,1723.
Antirestriction systems such as antirestriction proteins
control RM systems.
Antirestriction systems such as antirestriction proteins
control RM systems.
ArdB from a
B. pertussis
plasmid
Ocr from phage T7
ArdA from transposon Tn916
Walkinshaw et al. Mol. Cell (2002) 9, 187. McMahon et al., (2009) Nucl. Acids Res. 37, 4887.
Serfiotis-Mitsa et al., (2010) Nucl. Acids Res. 38, 1723.
Bacterial DNA restriction and modification systems: structure,
function and evolution.
1.
Restriction and modification (RM) systems and their inhibition by
antirestriction systems.
2.
Type I RM enzymes: structure, function and evolution.
3.
Some speculation on the antiquity of RM systems and their
influence on evolution.
The components of Type I DNA RM enzymes.
Type I RM enzyme
comprises 5 subunits:
R2M2S1
Highly conserved in sequence and
structure except for target recognition
domains in the S subunits.
Molecular mass:
440,000
Kennaway et al. Genes and Development (2012) 26, 92-104.
Type I RM systems in S. aureus allow changes of specificity.
HsdS
CC group /
enzyme
Trd1
spacer
Trd2
CC1-1
CCAY
(N)5
TTAA
CC1-2
CCAY
(N)6
TGT
CC5-1
ATC
(N)5
CCT
CC5-2
CCAY
(N)6
GTA
CC133-771
CAG
(N)5
RTGA
CC398-1
ACC
(N)5
RTGA
26 other CC
12 others
14 others
EM model for a Type I RM enzyme with DNA bound. HsdR (red),
HsdM (blue and cyan), HsdS (yellow).
Kennaway et al. Genes and Development (2012) 26, 92-104.
Type I RM systems in S. aureus control horizontal gene transfer.
promoter
R
N-term
Nuclease
Motor
tail
CC1, CC5,
CC8/ST239
HsdR
promoter
M
frameshift
N-term
catalytic
tail
S
cr
promoter
M
N-term
catalytic
cr
CC1-1
cr
CCAY
6
TGT
cr
CC1-2 and CC8/ST239
5
CCT
cr
CC5-1 and CC8/ST239
6
GTA
cr
CC5-2
frameshift
N-term
catalytic
tail
S
cr
promoter
ATC
frameshift
N-term
catalytic
tail
S
HsdM
TTAA
tail
promoter
M
5
frameshift
S
M
CCAY
cr
CCAY
HsdS
Roberts et al. Nucl. Acids Res.
doi: 10.1093/nar/gkt535
Bacterial DNA restriction and modification systems: structure,
function and evolution.
1.
Restriction and modification (RM) systems and their inhibition by
antirestriction systems.
2.
Type I RM enzymes: structure, function and evolution.
3.
Some speculation on the antiquity of RM systems and
their influence on evolution.
What was the first RM system?
What did the first RM
system look like?
When did it appear?
What pressure caused it to
diversify? (Antirestriction
perhaps?)
Williams. J. Inorg. Biochem. (2006), 100, 1908
Why was the first RM system not a Type II RM system?
They are too specialised.
Type II restriction enzymes highly variable in sequence and structure but the methylases
are conserved.
Pingoud and Jeltsch. Nucl Acids
Res (2001) 29, 3705
Cheng and Roberts. Nucl Acids
Res (2001) 29, 3784
Subtypes of Type II restriction enzymes.
What was the first RM system?
Perhaps it was a proto-Type I RM enzyme?
The first RM system could be built from
DNA repair enzymes, small-molecule
methylases and transcription factors.
These would be present before the
LUCA.
Most antirestriction targets Type I RM
systems. This would force diversification
in RM structure to avoid antirestriction
DNA mimics and create new RM Types.
What would happen to Horizontal Gene Transfer
with "perfect" restriction and modification systems
or "perfect" antirestriction?
Why did RM and
anti-RM appear?