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7.1 Major Modes of Gene 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 by fine tuning protein levels © 2015 Pearson Education, Inc. Levels of Regulation A Snapshot –35 –10 +1 Promoter RBS Structural gene Terminator DNA 5′ 3′ 5′ 3′ Activation Repression Transcription (making RNA) RBS RNA Start codon Stop codon 5′ 3′ 5′-UTR 3′-UTR Translation (making protein) Feedback inhibition Mechanisms of controlling enzyme activity Protein Degradation © 2015 Pearson Education, Inc. Protein–protein interactions Covalent modifications Figure 7.1 II. DNA-Binding Proteins and Transcriptional Regulation • 7.2 DNA-Binding Proteins • 7.3 Negative Control: Repression and Induction • 7.4 Positive Control: Activation • 7.5 Global Control and the lac Operon • 7.6 Transcriptional Controls in Archaea © 2015 Pearson Education, Inc. 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 © 2015 Pearson Education, Inc. 7.2 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 © 2015 Pearson Education, Inc. 7.2 DNA-Binding Proteins • Homodimeric proteins: proteins composed of two identical polypeptides • Protein dimers interact with inverted repeats on DNA © 2015 Pearson Education, Inc. Types of DNA Repeats © 2015 Pearson Education, Inc. Mirror repeat 7.2 DNA-Binding Proteins • Several classes of protein domains are critical for proper binding of proteins to DNA • Helix-turn-helix (Figure 7.4)-one class of DNA binding domain • First helix is the recognition helix • Second helix is the stabilizing helix © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. A perfect fit for the major groove © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. 7.2 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 © 2015 Pearson Education, Inc. Three important types of DNA binding domains © 2015 Pearson Education, Inc. 7.3 Negative Control: Repression and Induction • Defined as control that prevents transcription • Several mechanisms in bacteria • These systems are greatly influenced by environment in which the organism is growing • Through presence or absence of specific small molecules • Interactions between small molecules and DNA-binding proteins result in control of transcription or translation © 2015 Pearson Education, Inc. 7.3 Negative Control: Repression and Induction • Early on, microbiologists realized that bacteria could respond to environmental signals by starting or stopping to make enzymes: adaptation. • Induction-occurs when environmental signal triggers the synthesis of an enzyme • Repression-occurs when environmental signal prevents the synthesis of an enzyme © 2015 Pearson Education, Inc. 7.3 Negative Control: Repression and Induction 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 © 2015 Pearson Education, Inc. Induction Total protein Cell number β-Galactosidase Lactose added © 2015 Pearson Education, Inc. Figure 7.6 7.3 Negative Control: Repression and Induction • 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) © 2015 Pearson Education, Inc. Repression Cell number Total protein Arginine added Arginine biosynthesis enzymes © 2015 Pearson Education, Inc. Figure 7.5 7.3 Negative Control: Repression and Induction • Inducer: a substance that induces enzyme synthesis • Corepressor: a substance that represses enzyme synthesis (not same as repressor) • Effectors: collective term for inducers and repressors • Effectors affect transcription indirectly by binding to specific DNA-binding proteins © 2015 Pearson Education, Inc. 7.3 Negative Control: Repression and Induction • Paradigm system is the lactose (lac) operon of E. coli. • Operon: group of genes related in that they work together • Structural genes code for enzymes or soldier proteins • Regulatory genes control the structural genes (Figure 7.7) • Enzyme induction can also be controlled by a repressor • Addition of inducer inactivates a repressor protein (not same as corepressor substance) , and transcription can proceed (Figure 7.8) • Repressor's role is to prevent enzyme synthesis, so it is called negative control © 2015 Pearson Education, Inc. 7.3 Negative Control: Repression and Induction • Structural genes of lac operon: (contiguous) • Lac Z • Lac Y • Lac A • Regulatory genes of the lac operon: (contiguous, overlapping or separate) • Lac operator or Lac O • Lac promoter or Lac P • Lac repressor or Lac I-coded by an independent gene with its own promoter-always active at a low level: constitutive © 2015 Pearson Education, Inc. lac Promoter lac Operator RNA polymerase lacZ lacY lacA Transcription blocked Repressor lac Promoter lac Operator RNA polymerase lacZ lacY lacA Transcription proceeds Repressor Inducer (allolactose) © 2015 Pearson Education, Inc. Figure 7.8 7.3 Negative Control: Repression and Induction • Structural genes of arg operon: (contiguous) • Arg C • Arg B • Arg H • Regulatory genes of the lac operon: (contiguous, overlapping or separate) • Arg operator or Arg O • Arg promoter or Arg P • Arg repressor or Arg R-coded by an independent gene with its own promoter-always active at a low level © 2015 Pearson Education, Inc. arg Promoter arg Operator argC argB RNA polymerase argH Transcription proceeds Repressor arg Promoter arg Operator RNA polymerase © 2015 Pearson Education, Inc. argC argB argH Corepressor Transcription blocked (arginine) Repressor Figure 7.7 Summary: Induction and repression work by way of a protein that changes shape in response to a signal © 2015 Pearson Education, Inc. 7.4 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 © 2015 Pearson Education, Inc. Activatorbinding site mal Promoter malE malF malG No transcription RNA polymerase Maltose activator protein Activatorbinding site mal Promoter malE RNA polymerase malF malG Transcription proceeds Maltose activator protein Inducer (maltose) © 2015 Pearson Education, Inc. Figure 7.9 7.4 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) © 2015 Pearson Education, Inc. Activatorbinding site Promoter RNA polymerase Transcription proceeds Activator protein Promoter RNA polymerase Activator protein Transcription proceeds Activatorbinding site © 2015 Pearson Education, Inc. Figure 7.11 7.4 Positive Control: Activation • Genes for maltose are spread out over the chromosome in several operons (Figure 7.12) • Each operon has an activator-binding site • Multiple operons controlled by the same regulatory protein are called a regulon • Regulons also exist for negatively controlled systems © 2015 Pearson Education, Inc. GF B EK M A Y Z oriC Mal regulatory protein Lac regulatory protein T P Q Maltose operons make up maltose regulon Lactose operon Direction of transcription © 2015 Pearson Education, Inc. Figure 7.12 7.5 Global Control and the lac Operon • Global control systems: regulate expression of many different genes simultaneously • Catabolite repression is an example of global control • Synthesis of unrelated catabolic enzymes is repressed if glucose is present in growth medium (Figure 7.13) • lac operon is under control of catabolite repression • Ensures that the "best" carbon and energy source is used first • Diauxic growth: two exponential growth phases © 2015 Pearson Education, Inc. Growth on lactose Glucose exhausted Growth on glucose © 2015 Pearson Education, Inc. Figure 7.13 7.5 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 7.15) • Cyclic AMP receptor protein (CRP) is the activator protein • Cyclic AMP is a key molecule in many metabolic control systems • Derived from a nucleic acid precursor • Is a regulatory nucleotide © 2015 Pearson Education, Inc. CRP protein cAMP RNA polymerase Binding of CRP recruits RNA polymerase DNA lacI Transcription mRNA lacI lac Structural genes C P O lacZ Active repressor binds to operator and blocks transcription. lacY lacA Transcription mRNA lacZ lacY lacA Translation Translation LacI Inducer LacZ Active repressor LacY LacA Lactose catabolism © 2015 Pearson Education, Inc. Inactive repressor Figure 7.15 7.5 Global Control and the lac Operon • Dozens of catabolic operons are 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 © 2015 Pearson Education, Inc. 7.6 Transcription Controls in Archaea • Archaea use DNA-binding proteins to control transcription • More closely resembles control by Bacteria than Eukarya • Repressor proteins in Archaea • NrpR is an example of an archaeal repressor protein from Methanococcus maripaludis (Figure 7.16) • Represses genes involved in nitrogen metabolism © 2015 Pearson Education, Inc. NrpR DNA NrpR blocks TFB and TBP binding; no transcription. BRE TATA INIT NrpR binds α-ketoglutarate. α-Ketoglutarate ( ) NH3 When NrpR is released, TBP and TFB can bind. Glutamate NrpR TFB TBP Transcription proceeds. RNA polymerase © 2015 Pearson Education, Inc. Figure 7.16 III. Sensing and Signal Transduction • 7.7 Two-Component Regulatory Systems • 7.8 Regulation of Chemotaxis • 7.9 Quorum Sensing • 7.10 Other Global Control Networks © 2015 Pearson Education, Inc. 7.7 Two-Component Regulatory Systems • Prokaryotes regulate cellular metabolism in response to environmental fluctuations • External signal is transmitted directly to the target • External signal is detected by sensor and transmitted to regulatory machinery (signal transduction) • Most signal transduction systems are two-component regulatory systems © 2015 Pearson Education, Inc. 7.7 Two-Component Regulatory Systems • Two-component regulatory systems (Figure 7.17) • 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 © 2015 Pearson Education, Inc. Environmental signal Sensor kinase ATP Cytoplasmic membrane ADP His His P P Response regulator Phosphatase activity P P RNA polymerase Promoter Transcription blocked Operator DNA Structural genes Shows regulator blocking transcription but activation can also occur © 2015 Pearson Education, Inc. 7.7 Two-Component Regulatory Systems • Almost 50 different two-component systems in E. coli • Examples include phosphate assimilation, nitrogen metabolism, and osmotic pressure response (Figure 7.18) • Some Archaea also have two-component regulatory systems • Some signal transduction systems have multiple regulatory elements © 2015 Pearson Education, Inc. 7.8 Regulation of Chemotaxis • Modified two-component system used in chemotaxis to • Sense temporal changes in attractants or repellents • Regulate flagellar rotation • Three main steps (Figure 7.19) 1. Response to signal 2. Controlling flagellar rotation 3. Adaptation © 2015 Pearson Education, Inc. 7.8 Regulation of Chemotaxis • Step 1: Response to signal • Sensory proteins in cytoplasmic membrane sense attractants and repellents • Methyl-accepting chemotaxis proteins (MCPs) • Bind attractant or repellent and initiate flagellar rotation • Step 2: Controlling flagellar rotation • Controlled by CheY protein • CheY results in counterclockwise rotation and runs • CheY-P results in clockwise rotation and tumbling © 2015 Pearson Education, Inc. Repellents bind to MCP and trigger phosphorylation of CheA-CheW complex. MCP CheR MCP is both methylated and demethylated. CheB P CheW CheA ATP ADP CheY CheA-CheW phosphorylate CheY and CheB. Flagellar motor CheY-P binds to flagellar switch. CheY P CheZ CheB Cytoplasm CheZ dephosphorylates CheY-P. Flagellum © 2015 Pearson Education, Inc. Figure 7.19 Summary • CheY = counterclockwise rotation = run • But CheY-P = clockwise = tumble • Repellents increase CheY-P therefore tumbling and direction change • Works through two component system using MCP and other Che proteins © 2015 Pearson Education, Inc. 7.8 Regulation of Chemotaxis • Step 3: Adaptation • Feedback loop • Allows the system to reset itself to continue to sense the presence of a signal • Involves modification of MCPs with methyl group • Degree of methylation controls sensitivity to attractant and repellent © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. 7.9 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 that a sufficient number of cells are present before initiating a response that, to be effective, requires a certain cell density (e.g., toxin production in pathogenic bacterium) © 2015 Pearson Education, Inc. 7.9 Quorum Sensing • Each species of bacterium produces a specific autoinducer molecule (Figure 7.20) • 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 © 2015 Pearson Education, Inc. Acyl homoserine lactone (AHL) Activator protein AHL AHL Quorumspecific proteins Other cells of the same species Chromosome © 2015 Pearson Education, Inc. AHL synthase Figure 7.20 7.9 Quorum Sensing • Several different classes of autoinducers • Acyl homoserine lactone (AHL) was the first autoinducer to be identified • Quorum sensing first discovered as mechanism regulating light production in bacteria including Aliivibrio fischeri (Figure 7.21) • Lux operon encodes bioluminescence © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. 7.9 Quorum Sensing • Examples of quorum sensing • Virulence factors • Switching from free-living to growing as a biofilm • Quorum sensing is present in some microbial eukaryotes • Quorum sensing likely exists in Archaea © 2015 Pearson Education, Inc. 7.9 Quorum Sensing-example • Virulence factors • Staphylococcus aureus • Secretes small peptides that damage host cells or alter host's immune system • Under control of autoinducing peptide (AIP) • Activates several proteins that lead to production of virulence proteins (Figure 7.22b) © 2015 Pearson Education, Inc. Binding of AIP to ArgC leads to auto-phosphorylation. AIP Cytoplasmic membrane ArgB ArgC ATP P Pre-AIP Cytoplasm ADP ArgC phosphorylates ArgA. Pre-AIP is converted to AIP by ArgB and exported out of the cell. ArgA P ArgA-P activates expression of genes required for pre-AIP and virulence proteins. + + Basal transcription D C B argA Virulence proteins Genes encoding virulence Virulence factor production in Staphylococcus © 2015 Pearson Education, Inc. Figure 7.22b Summary • Staphylococcus aureus is an opportunitist pathogen • ArgA-P activates arg genes above basal level • Also activates virulence factors Exoenzymes to digest tissue Exotoxins as poisons (gastroenteritis) Role in toxic shock syndrome (TSS) © 2015 Pearson Education, Inc. 7.9 Quorum Sensing • Biofilm formation • Pseudomonas aeruginosa • Produces polysaccharides that increase pathogenicity and antibiotic resistance • Two quorum-sensing systems • Produces AHLs and cyclic di-guanosine monophosphate (c-di-GMP) • Leads to exopolysaccharide production and flagella synthesis (Figure 7.23) © 2015 Pearson Education, Inc. Production of AHLs and c-di-GMP Increasing cell population © 2015 Pearson Education, Inc. Exopolysaccharide production and flagella synthesis Attachment Mature biofilm Figure 7.23 7.10 Other Global Control Networks • Nitrogen utilization: regulation by alternate sigma • Under limiting conditions nitrogen uptake and use becomes very important • Requires expression of new genes • The genes do not have consensus promoter sequence, therefore not recognized by regular RNA pol and sigma factor • Nitrogen stress results in expression of alternate sigma: sigma54 from RpoN gene © 2015 Pearson Education, Inc. 7.13 Nitrogen Fixation, Nitrogenase, and Heterocyst Formation • Nitrogen fixation is process of reducing N2 to NH3 • Only certain prokaryotes can fix nitrogen • Reaction is catalyzed by nitrogenase • Composed of dinitrogenase and dinitrogenase reductase • Sensitive to the presence of oxygen © 2015 Pearson Education, Inc. 7.13 Nitrogen Fixation, Nitrogenase, and Heterocyst Formation • Highly regulated process because it is such an energy-demanding process • Nif regulon coordinates regulation of genes essential to nitrogen fixation (Figure 7.27) • Oxygen and ammonia are the two main regulatory effectors • Complex regulation uses multiple strategies © 2015 Pearson Education, Inc. Nitrogenase proteins Dinitrogenase reductase FeMo-co synthesis Mo processing FeMo-co synthesis Dinitrogenase Dinitrogenase Homocitrate reductase synthesis processing Regulators Metal center Positive Negative biosynthesis Flavodoxin FeMo-co synthesis β Electron transport Pyruvate flavodoxin oxidoreductase α FeMo-co insertion into dinitrogenase nif DNA Q B A L F M Z W V S U X N E Y T K D H J RNA © 2015 Pearson Education, Inc. Figure 7.27 7.13 Nitrogen Fixation, Nitrogenase, and Heterocyst Formation • Heterocyst-specialized cells for nitrogen fixation in some cyanobacteria (Figure 7.28) • Requires metabolic and morphological changes • Formation of thickened envelope (3 layers) • Inactivation of photosystem II (it releases oxygen) • Expression of nitrogenase • Patterning of heterocyst differentiation © 2015 Pearson Education, Inc. Fixed N flow [α-Ketoglutarate] NtcA activates hetR expression Heterocyst Vegetative cells Vegetative cells Fixed C flow A filament of Anabaena © 2015 Pearson Education, Inc. Heterocyst—vegetative cell interactions HetR activates genes necessary for heterocyst formation Triggering heterocyst formation Figure 7.28 V. RNA-Based Regulation • Uses non-coding RNAs or non-coding regions of transcripts for regulation • 7.14 Regulatory RNAs: Small RNAs (i.e. antisense or cis-RNA and trans-RNA) • 7.15 Riboswitches • 7.16 Attenuation © 2015 Pearson Education, Inc. 7.14 Regulatory RNA: Small RNAs • Small RNAs work by: • Block or open a ribosome-binding site (RBS) • Increase or decrease degradation of mRNA (i.e. mRNA half-life) • May also act at level of transcription © 2015 Pearson Education, Inc. Translation inhibition/stimulation 1. mRNA 5′ 3′ 3′ sRNA RNA degradation/protection 1. 5′ 5′ 3′ RBS mRNA 5′ RBS 3′ 3′ sRNA 5′ RBS 5′ 3′ RBS Ribonuclease Translation No translation Translation No translation 3′ 2. 5′ RBS 2. 5′ 3′ 5′ 3′ RBS 5′ 3′ RBS 3′ 5′ 5′ 3′ RBS Ribonuclease No translation © 2015 Pearson Education, Inc. Translation No translation Translation Figure 7.29 7.14 Regulatory RNA: Small RNAs • Antisense RNAs first discovered in plasmids, phages, transposons • RNA-OUT of Tn10 (IS10) decreases transposase expression • Most antisense regulators have not been studied • How do we know they exist? © 2015 Pearson Education, Inc. 7.14 Regulatory RNA: Small RNAs and Antisense RNA • Second classic example • AntiQ RNA from Enterococcus faecalis plasmid pCF10 • Small RNA binds to growing mRNA transcript for conjugation genes • Alters secondary structure to block further transcription © 2015 Pearson Education, Inc. 7.14 Regulatory RNA: Small RNAs and Antisense RNA • Types of small RNAs (cont'd) • Trans-RNAs are encoded in the intergenic region (not within gene they regulate) • Limited complementarity with target • Usually require help from a protein for binding © 2015 Pearson Education, Inc. 7.14 Regulatory RNA: Small RNAs and Antisense RNA • Hfq protein • Facilitates RNA binding • Also has regulatory functions independent of RNA • “Global” regulator • http://www.rcsb.org/pdb/pv/pv.do?pdbid=1K Q2&bionumber=1 © 2015 Pearson Education, Inc. Small regulatory RNA mRNA 5′ Hfq protein © 2015 Pearson Education, Inc. 3′ Small regulatory RNA recognition sequence Figure 7.30 Useful Reference •Annu Rev Genet. Author manuscript; available in PMC 2011 Jan 28. •Annu Rev Genet. 2010; 44: 167–188. •doi: 10.1146/annurev-genet-102209-163523 •PMCID: PMC3030471 •NIHMSID: NIHMS236247 •Bacterial antisense RNAs: How many are there and what are they doing? •Maureen Kiley Thomason1,2 and Gisela Storz1 •1 Cell Biology and Metabolism Program, Eunice Kennedy Shriver National Institute of Child Health and Human Development, Bethesda, MD 20892-5430 •2 Department of Biochemistry and Molecular & Cell Biology, Georgetown University Medical Center, Washington, DC 20007 •Maureen Kiley Thomason: vog.hin.liam@amyelik; Gisela Storz: vog.hin.xileh@zrots •Abstract •Antisense RNAs encoded on the DNA strand opposite another gene have the potential to form extensive base pairing interactions with the corresponding sense RNA. Unlike other smaller regulatory RNAs in bacteria, antisense RNAs range in size, from tens to thousands of nucleotides. The numbers of antisense RNAs reported for different bacteria vary extensively but hundreds have been suggested in some species. If all of these reported antisense RNAs are expressed at levels sufficient to regulate the genes encoded opposite them, antisense RNAs could significantly impact gene expression in bacteria. Here we review the evidence for these RNA regulators and describe what is known about the functions and mechanisms of action for some of these RNAs. Important considerations for future research as well as potential applications are also discussed. © 2015 Pearson Education, Inc. 7.15 Riboswitches • Riboswitches: RNA domains in an mRNA molecule that can bind small molecules to control translation of mRNA (Figure 7.31) • Located at 5′ end of mRNA • Binding results from folding of RNA into a 3-D structure • Similar to a protein recognizing a substrate • Found in some bacteria, fungi, and plants © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Figure 7.31 7.15 Riboswitches • Riboswitch example: SAM riboswitch or (SAM Box riboswitch) in Bacillus subtilis • Regulates expression of genes required for methionine metabolism at level of translation of mRNA No SAM: transcription of mRNA takes place SAM causes shape change that allows formation of a terminator © 2015 Pearson Education, Inc. 7.16 Attenuation • Transcriptional control that functions by premature termination of mRNA synthesis • Trp operon of E. coli • Additional control level above and beyond induction or repression • Presence of trp down regulates expression of genes for trp biosynthesis • Depends on control sequences in 5’UTR or trp leader © 2015 Pearson Education, Inc. trp structural genes P O L DNA trpE trpD trpC trpB trpA Trp Leader Met-Lys-Ala-lle-Phe-Val-Leu-Lys-Gly-Trp-Trp-Arg-Thr-Ser Trp mRNA gets translated as it is made When there is trp in the cell the trp leader can be transcribed No trp in the cell transcription stalls © 2015 Pearson Education, Inc. Figure 7.32 Excess tryptophan: transcription terminated Leader sequence DNA Direction of transcription Base pairing Ribosome 2 5′ 3 1 Trp-rich leader peptide Transcription terminated and tryptophan structural genes not transcribed Direction of translation Leader sequence DNA Direction of transcription Translation stalled 2 5′ 1 Direction of translation © 2015 Pearson Education, Inc. 4 mRNA Limiting tryptophan: transcription proceeds Leader peptide RNA polymerase terminates 3 RNA polymerase continues 4 Transcription continues and tryptophan structural genes transcribed Figure 7.33 VI. Regulation of Enzymes and Other Proteins Post-translational • 7.17 Feedback Inhibition • 7.18 Post-Translational Regulation © 2015 Pearson Education, Inc. 7.17 Feedback Inhibition • Feedback inhibition: mechanism for turning off the reactions in a biosynthetic pathway (Figure 7.34a) • End product of the pathway binds to the first enzyme in the pathway, thus inhibiting its activity • Inhibited enzyme is an allosteric enzyme (Figure 7.34b) • Two binding sites: active and allosteric • Easily reversible reaction • Fast-acting © 2015 Pearson Education, Inc. 7.17 Feedback Inhibition • Some pathways controlled by feedback inhibition use isoenzymes, different enzymes that catalyze the same reaction but are subject to different regulatory controls • Rhodopseudomonas palustris • Highly versatile bacterium with multiple life styles • Switches between different nitrogenase isoenzymes depending on conditions © 2015 Pearson Education, Inc. 7.18 Post-Translational Covalent Modification • Biosynthetic enzymes can also be regulated by covalent modifications • Regulation involves a small molecule attached to or removed from the protein (Figure 7.35) • Results in conformational change that inhibits activity • Common modifiers include adenosine monophosphate (AMP), adenosine diphosphate (ADP), inorganic phosphate (PO42-), and methyl groups (CH3) © 2015 Pearson Education, Inc.