Bacteriology course
Jeong Nam Kim
Department of Microbiology
Pusan National University
1

Major Modes of Regulation
• Gene expression: transcription of gene into mRNA followed by translation of mRNA into protein (Figure 7.1)
• Most proteins are enzymes that carry out biochemical reactions
• Constitutive proteins are needed at the same level all the time
• Microbial genomes encode many proteins that are not needed all the time
• Regulation helps conserve energy and resources

Major Modes of Regulation
• Two major levels of regulation in the cell:
– One controls the activity of preexisting enzymes
• Post-translational regulation
• Very rapid process (seconds)
– One controls the amount of an enzyme
• Regulates level of transcription
• Regulates translation
• Slower process (minutes)

Major Modes of Regulation
• Monitoring gene expression
– Reporter genes encode for an easy-to-detect product
(Figure 7.2)
• Can be fused to other genes
• Can be fused to regulatory elements
• Example: green fluorescent protein (GFP)

Figure 7.2

DNA-Binding Proteins
• mRNA transcripts generally have a short half-life
– Prevents the production of unneeded proteins
• Regulation of transcription typically requires proteins that can bind to DNA
• Small molecules influence the binding of regulatory proteins to DNA
– Proteins actually regulate transcription

DNA-Binding Proteins
• Most DNA-binding proteins interact with DNA in a sequencespecific manner
• Specificity provided by interactions between amino acid side chains and chemical groups on the bases and sugar–phosphate backbone of DNA
• Major groove of DNA is the main site of protein binding
• Inverted repeats frequently are binding site for regulatory proteins

DNA-Binding Proteins
• Homodimeric proteins: proteins composed of two identical polypeptides
• Protein dimers interact with inverted repeats on DNA
– Each of the polypeptides binds to one inverted repeat (Figure 7.3)

Domain containing protein–protein contacts, holding protein dimer together
DNA-binding domain fitsin major grooves and along sugar–phosphate backbone
Inverted repeats
5 ′
3 ′
TG T G T G G A A T TGT GA GC GGA T A A C A A T T T C A C A C A 3 ′
AC AC A C C T T AACA C T C GC C T A T T GT T A AA G T G T G T 5 ′
Inverted repeats
Figure 7.3

DNA-Binding Proteins
• Several classes of protein domains are critical for proper binding of proteins to DNA
– Helix-turn-helix (Figure 7.4)
• First helix is the recognition helix
• Second helix is the stabilizing helix
• Many different DNA-binding proteins from Bacteria contain helix-turn-helix
– lac and trp repressors of E. coli

Recognition helix
Turn
Stabilizing helix
DNA
Subunits of binding protein
Figure 7.4

DNA-Binding Proteins
• Classes of protein domains
– Zinc finger
• Protein structure that binds a zinc ion
• Eukaryotic regulatory proteins use zinc fingers for DNA binding
– Leucine zipper
• Contains regularly spaced leucine residues
• Function is to hold two recognition helices in the correct orientation

DNA-Binding Proteins

DNA-Binding Proteins
• Multiple outcomes after DNA binding are possible
1. DNA-binding protein may catalyze a specific reaction on the DNA molecule (i.e., transcription by RNA polymerase)
2. The binding event can block transcription
(negative regulation)
3. The binding event can activate transcription
(positive regulation)

Negative Control: Repression and
Induction
• Several mechanisms for controlling gene expression in bacteria
– These systems are greatly influenced by environment in which the organism is growing
– Presence or absence of specific small molecules
– Interactions between small molecules and DNAbinding proteins result in control of transcription or translation

Negative Control: Repression and
Induction
• Negative control: a regulatory mechanism that stops transcription
– Repression: preventing the synthesis of an enzyme in response to a signal (Figure 7.5)
• Enzymes affected by repression make up a small fraction of total proteins
• Typically affects anabolic enzymes
(e.g., arginine biosynthesis)

Repression
Cell number
Total protein
Arginine added
Arginine biosynthesis enzymes
Figure 7.5

Negative Control: Repression and
Induction
• Negative control (cont'd)
– Induction: production of an enzyme in response to a signal (Figure 7.6)
• Typically affects catabolic enzymes (e.g., lac operon)
• Enzymes are synthesized only when they are needed
– No wasted energy

Induction
Total protein
Cell number
β-Galactosidase
Lactose added
Figure 7.6

Negative Control: Repression and
Induction
• Inducer: substance that induces enzyme synthesis
• Corepressor: substance that represses enzyme synthesis
• Effectors: collective term for inducers and repressors
• Effectors affect transcription indirectly by binding to specific
DNA-binding proteins
– Repressor molecules bind to an allosteric repressor protein
– Allosteric repressor becomes active and binds to region of DNA near promoter called the operator

• Operon: cluster of genes arranged in a linear fashion whose expression is under control of a single operator
– Operator is located downstream of the promoter
– Transcription is physically blocked when repressor binds to operator
(Figure 7.7)
• Enzyme induction can also be controlled by a repressor
– Addition of inducer inactivates repressor, and transcription can proceed
(Figure 7.8)
• Repressor's role is inhibitory, so it is called negative control

arg Promoter arg Operator argC argB
RNA polymerase argH
Transcription proceeds
Repressor
arg Promoter arg Operator argC argB
RNA polymerase
Corepressor
(arginine)
Repressor argH
Transcription blocked
Figure 7.7

lac Promoter lac Operator lacZ
RNA polymerase
Repressor lacY lacA
Transcription blocked
lac Promoter lac Operator lacZ
RNA polymerase lacY lacA
Transcription proceeds
Repressor
Inducer
(allolactose)
Figure 7.8

Positive Control: Activation
• Positive control: regulator protein activates the binding of RNA polymerase to DNA (Figure
7.9)
• Maltose catabolism in E. coli
– Maltose activator protein cannot bind to DNA unless it first binds maltose
• Activator proteins bind specifically to certain
DNA sequence
– Called activator-binding site, not operator

Activatorbinding site
mal Promoter malE
RNA polymerase
Maltose activator protein malF malG
No transcription
Activatorbinding site
mal Promoter malE
RNA polymerase
Maltose activator protein
Inducer
(maltose) malF malG
Transcription proceeds
Figure 7.9

Positive Control: Activation
• Promoters of positively controlled operons only weakly bind
RNA polymerase
• Activator protein helps RNA polymerase recognize promoter
– May cause a change in DNA structure
– May interact directly with RNA polymerase
• Activator-binding site may be close to the promoter or be several hundred base pairs away (Figure 7.11)

Activatorbinding site
Promoter
RNA polymerase
Activator protein
Transcription proceeds
Activator protein
Promoter
RNA polymerase
Transcription proceeds
Activatorbinding site
Figure 7.11