8.1 Major Modes of Regulation Gene expression: transcription of gene into mRNA followed by...
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8.1 Major Modes of Regulation
• Gene expression: transcription of gene into mRNA followed by translation of mRNA into protein (Figure 8.1)
• 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
© 2012 Pearson Education, Inc.
Figure 8.1 Upstreamregion
Downstreamregion
Start oftranscription Shine-Dalgarno
sequence (ribosome-binding site)
Start codon:Translationstarts here
Stop codon:Translation ends here
Transcriptionterminator
Transcription
Translation
Protein
DNA
mRNA
© 2012 Pearson Education, Inc.
8.2 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
© 2012 Pearson Education, Inc.
8.2 DNA-Binding Proteins
• Most DNA-binding proteins interact with DNA in a sequence-specific 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
© 2012 Pearson Education, Inc.
8.2 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 8.2)
© 2012 Pearson Education, Inc.
Figure 8.2
Domain containing protein–proteincontacts, holding protein dimer together
DNA-binding domain fits inmajor grooves and alongsugar–phosphate backbone
Inverted repeats
Inverted repeats
© 2012 Pearson Education, Inc.
8.2 DNA-Binding Proteins
• Several classes of protein domains are critical for proper binding of proteins to DNA
– Helix-turn-helix (Figure 8.3)• 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
© 2012 Pearson Education, Inc.
Figure 8.3
Stabilizinghelix
Recognitionhelix
Subunitsof bindingprotein
Turn
DNA
© 2012 Pearson Education, Inc.
8.2 DNA-Binding Proteins
• Multiple outcomes after DNA binding are possible1. 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)
© 2012 Pearson Education, Inc.
8.3 Negative Control of Transcription: 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 DNA-binding proteins result in control of transcription or translation
© 2012 Pearson Education, Inc.
• Negative control: a regulatory mechanism that stops transcription– Repression: preventing the synthesis of an
enzyme in response to a signal (Figure 8.5)• Enzymes affected by repression make up a
small fraction of total proteins • Typically affects anabolic enzymes
(e.g., arginine biosynthesis)
8.3 Negative Control of Transcription: Repression and Induction
© 2012 Pearson Education, Inc.
8.3 Negative Control of Transcription: Repression and Induction
• Negative Control (cont’d)– Induction: production of an enzyme in response to
a signal (Figure 8.6)• Typically affects catabolic enzymes (e.g., lac
operon)• Enzymes are synthesized only when they are
needed– no wasted energy
© 2012 Pearson Education, Inc.
Figure 8.5
Repression
Cell numberTotal protein
Arginine added
Argininebiosynthesisenzymes
Time
Re
lati
ve
inc
rea
se
© 2012 Pearson Education, Inc.
Figure 8.6
Induction
Cell number
Total protein
Lactose added
-Galacto-sidase
Time
Re
lati
ve
inc
rea
se
© 2012 Pearson Education, Inc.
8.3 Negative Control of Transcription: 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© 2012 Pearson Education, Inc.
Figure 8.7
RNApolymerase
Repressor
Repressor
RNApolymerase
Corepressor(arginine)
Transcription proceeds
Transcription blocked
arg Promoter arg Operator argC argB argH
arg Promoter arg Operator argC argB argH
© 2012 Pearson Education, Inc.
Figure 8.8
RNApolymerase
Repressor
Repressor
RNApolymerase
Inducer
Transcription proceeds
Transcription blocked
lac Promoter lac Operator lacZ lacY lacA
lac Promoter lac Operator lacZ lacY lacA
© 2012 Pearson Education, Inc.
8.4 Positive Control of Transcription
• Positive control: regulator protein activates the binding of RNA polymerase to DNA (Figure 8.9)
• 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
© 2012 Pearson Education, Inc.
Figure 8.9
Activator-binding site
RNApolymerase
Transcription proceeds
mal Promoter malE malF malG
No transcription
Activator-binding site
Maltose activator protein
Maltose activator protein
Inducer
RNApolymerase
mal Promoter malE malF malG
© 2012 Pearson Education, Inc.
8.4 Positive Control of Transcription
• 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 several hundred base pairs away (Figure 8.11)
© 2012 Pearson Education, Inc.
Figure 8.11
Activator-binding site
Activatorprotein
Activator-binding site
Activator protein
RNApolymerase
RNApolymerase
Promoter
Promoter
Transcriptionproceeds
Transcriptionproceeds
© 2012 Pearson Education, Inc.
8.5 Global Control and the lac Operon
• Cyclic AMP and CRP– In catabolite repression, transcription is controlled
by an activator protein and is a form of positive control (Figure 8.14)
– Cyclic AMP receptor protein (CRP) is the activator protein
– Cyclic AMP is a key molecule in many metabolic control systems
• It is derived from a nucleic acid precursor• It is a regulatory nucleotide
© 2012 Pearson Education, Inc.
Figure 8.14 CRP protein
cAMP
RNApolymerase
lac Structural genes
Activerepressor
Inducer
Inactiverepressor
Lactose catabolism
DNA
mRNA mRNA
TranscriptionTranscription
TranslationTranslation
lacZ lacY lacAlacI
Activerepressorbinds tooperatorandblockstran-scription
© 2012 Pearson Education, Inc.
8.5 Global Control and the lac Operon
• Dozens of catabolic operons affected by catabolite repression– Enzymes for degrading lactose, maltose, and
other common carbon sources
• Flagellar genes are also controlled by catabolite repression– No need to swim in search of nutrients
© 2012 Pearson Education, Inc.
8.7 Two-Component Regulatory Systems
• Prokaryotes regulate cellular metabolism in response to environmental fluctuations– External signal is transmitted directly to the target
– External signal detected by sensor and transmitted to regulatory machinery (Signal transduction)
• Most signal transduction systems are two-component regulatory systems
© 2012 Pearson Education, Inc.
8.7 Two-Component Regulatory Systems
• Two-component regulatory systems (Figure 8.16)– Made up of two different proteins:
• Sensor kinase: (in cytoplasmic membrane) detects environmental signal and autophosphorylates
• Response regulator: (in cytoplasm) DNA-binding protein that regulates transcription
– Also has feedback loop• Terminates signal
© 2012 Pearson Education, Inc.
Figure 8.16Environmental signal
Phosphataseactivity
Transcription blockedRNApolymerase
Sensorkinase
Cytoplasmicmembrane
Response regulator
DNA
Promoter Operator Structural genes© 2012 Pearson Education, Inc.
8.7 Two-Component Regulatory Systems
• Almost 50 different two-component systems in E. coli– Examples include phosphate assimilation,
nitrogen metabolism, and osmotic pressure response
• Some signal transduction systems have multiple regulatory elements
• Some Archaea also have two-component regulatory systems
© 2012 Pearson Education, Inc.
8.9 Quorum Sensing
• Prokaryotes can respond to the presence of other cells of the same species
• Quorum sensing: mechanism by which bacteria assess their population density
– Ensures sufficient number of cells are present before initiating a response that requires a certain cell density to have an effect (e.g., toxin production in pathogenic bacterium)
© 2012 Pearson Education, Inc.
8.9 Quorum Sensing
• Each species of bacterium produces a specific autoinducer molecule (Figure 8.18)– Diffuses freely across the cell envelope
– Reaches high concentrations inside cell only if many cells are near
– Binds to specific activator protein and triggers transcription of specific genes
© 2012 Pearson Education, Inc.
Figure 8.18
Acyl homoserine lactone (AHL)
AHLActivator protein
Other cellsof the samespecies
Chromosome AHL synthase
AHL
Quorum-specificproteins
© 2012 Pearson Education, Inc.
8.9 Quorum Sensing
• Several different classes of autoinducers– Acyl homoserine lactone was the first
autoinducer to be identified
• Quorum sensing first discovered as mechanism regulating light production in bacteria including Aliivibrio fischeri (Figure 8.19)
– Lux operon encodes bioluminescence
© 2012 Pearson Education, Inc.
Figure 8.19
© 2012 Pearson Education, Inc.
8.9 Quorum Sensing
• Examples of quorum sensing– P. aeruginosa switches from free living to
growing as a biofilm
– Virulence factors of Staphylococcus aureus
• Quorum sensing is present in some microbial eukaryotes
• Quorum sensing likely exists in Archaea
© 2012 Pearson Education, Inc.
8.11 Other Global Control Networks
• Several other global control systems – Aerobic and anaerobic respiration
– Catabolite repression
– Nitrogen utilization
– Oxidative stress
– SOS response
– Heat shock response
© 2012 Pearson Education, Inc.
8.11 Other Global Control Networks
• Heat shock response:– Largely controlled by alternative sigma factors
(Figure 8.21)
– Heat shock proteins: counteract damage of denatured proteins and help cell recover from temperature stress
• Very ancient proteins
• Heat shock response also occurs in Archaea
© 2012 Pearson Education, Inc.
Figure 8.21
RpoHDnaK
Degradation ofRpoH by protease
RpoH
RpoH isreleased
Proteins unfold athigh temperature
DnaK bindsunfoldedproteins
RpoH is freeto transcribeheat shockgenes
Hig
h t
em
pe
ratu
reL
ow
te
mp
era
ture
© 2012 Pearson Education, Inc.
IV. Regulation of Development in Model Bacteria
• 8.12 Sporulation in Bacillus• 8.13 Caulobacter Differentiation
© 2012 Pearson Education, Inc.
8.12 Sporulation in Bacillus
• Regulation of development in model bacteria– Some prokaryotes display the basic principle of
differentiation
• Endospore formation in Bacillus (Figure 8.22)– Controlled by 4 sigma factors
– Forms inside mother cell
– Triggered by adverse external conditions (i.e., starvation or desiccation)
© 2012 Pearson Education, Inc.