Microbial GeneticsMicrobial GeneticsThe Operon: The Operon:

Regulation of bacterial gene expressionRegulation of bacterial gene expression

Controlling enzymesControlling enzymesWe know that We know that

… … Bacterial cells carry out a huge number of Bacterial cells carry out a huge number of chemical reactions catalyzed by enzymeschemical reactions catalyzed by enzymes

And ….. Bacterial cells respond rapidly to And ….. Bacterial cells respond rapidly to changing environments e.g., presence of a changing environments e.g., presence of a new carbon sourcenew carbon source

So ….So ….

control of Enzymes is VERY important and control of Enzymes is VERY important and must be efficientmust be efficient

Enzyme activity can be regulated by feedbackEnzyme activity can be regulated by feedbackinhibition of synthesized enzymeinhibition of synthesized enzyme

BUTBUT making enzyme requires energy a a bacterium making enzyme requires energy a a bacterium

cannot make ALL enzymes ALLTHE TIMEcannot make ALL enzymes ALLTHE TIME**SOSO it is better to stop synthesizing enzymes that are not it is better to stop synthesizing enzymes that are not

neededneeded

THUS,THUS, many genes such as those for enzymes many genes such as those for enzymes involved in lactose metabolism or amino acid involved in lactose metabolism or amino acid production are regulated and only expressed production are regulated and only expressed when requiredwhen required

Enzyme Regulation

** However, 60-80% of genes are not regulated and are termed However, 60-80% of genes are not regulated and are termed ConstitutiveConstitutive

Regulation of gene expressionRegulation of gene expression

Constitutive Enzymes – No regulation

Enzyme Induced

Enzyme Repressed

Regulation of gene expressionRegulation of gene expression

The process of transcription plays an The process of transcription plays an important role in the ability of bacteria to important role in the ability of bacteria to respond to changing environments: mRNA respond to changing environments: mRNA compared to DNA is inherently unstable.compared to DNA is inherently unstable.

Thus, the expression levels of certain Thus, the expression levels of certain proteins can be controlled at the level of proteins can be controlled at the level of transcription (mRNA synthesis).transcription (mRNA synthesis).

Transcriptional controlTranscriptional control

Two genetic control mechanisms which Two genetic control mechanisms which regulate transcription of mRNA and, regulate transcription of mRNA and, therefore the synthesis of corresponding therefore the synthesis of corresponding proteinprotein

InductionInduction-turning on gene expression-turning on gene expression RepressionRepression-turning off gene expression-turning off gene expression

RepressionRepression Inhibition of gene expressionInhibition of gene expression Often in response to an Often in response to an

excess of endproduct, shuts excess of endproduct, shuts down synthesis of the down synthesis of the enzymeenzyme

Mediated by regulatory Mediated by regulatory proteins called proteins called repressorsrepressors Block RNA polymerase from Block RNA polymerase from

initiating transcriptioninitiating transcription Repressible genes are Repressible genes are

transcribed until they are transcribed until they are repressedrepressed

InductionInduction

Turns on transcription of a geneTurns on transcription of a gene A substance which acts to induce A substance which acts to induce

transcription of a gene is an transcription of a gene is an inducerinducer,, often the substrateoften the substrate

Enzymes synthesized in the presence of Enzymes synthesized in the presence of inducers are called inducers are called inducible enzymesinducible enzymes Example: lactose utilization genesExample: lactose utilization genes

Inducible genes are not transcribed until Inducible genes are not transcribed until they are inducedthey are induced

The operonThe operon

Multiple genes are arranged in the same Multiple genes are arranged in the same orientation and are closely linked on the DNAorientation and are closely linked on the DNA

Genes in an operon are transcribed on a single Genes in an operon are transcribed on a single RNA transcript, but are translated individually to RNA transcript, but are translated individually to form multiple proteins form multiple proteins A mechanism for coordinate control of genes involved A mechanism for coordinate control of genes involved

in a single process by regulating transcription of the in a single process by regulating transcription of the operon mRNAoperon mRNA

promoter operator

mRNA

proteins

The operon (cont…)The operon (cont…) The term “operon” not only includes the The term “operon” not only includes the

structural genes in the operon but also the structural genes in the operon but also the regulatory sequences controlling regulatory sequences controlling transcriptiontranscription PromoterPromoter-site of RNA polymerase binding-site of RNA polymerase binding OperatorOperator-site of binding of a regulatory protein-site of binding of a regulatory protein

promoter operator mRNA

Operons and regulationOperons and regulation

About 27% of About 27% of E. coliE. coli genes are in operons genes are in operons Many of the genes arranged in operons Many of the genes arranged in operons

are regulated are regulated For example,For example,

genes for the lactose utilization are controlled by genes for the lactose utilization are controlled by the the presence of lactosepresence of lactose

genes for pathogenesis are often controlled by genes for pathogenesis are often controlled by temperature or physiologic factortemperature or physiologic factor

Two regulated operonsTwo regulated operons

Two Two E. coliE. coli operons reflect how operons in operons reflect how operons in general are regulatedgeneral are regulated

The The laclac operon operon Encodes inducible catalytic enzymes involved Encodes inducible catalytic enzymes involved

in the lactose utilization and uptakein the lactose utilization and uptake The The argarg operon operon

Encodes repressible anabolic enzymes Encodes repressible anabolic enzymes involved in the production of the amino acid involved in the production of the amino acid argininearginine

The The laclac operon operon Three structural genesThree structural genes

lacZlacZ: : ββ-galactosidase: splits lactose to glucose and galactose-galactosidase: splits lactose to glucose and galactose lacYlacY: a permease involved in transport of lactose: a permease involved in transport of lactose lacAlacA: a transacetylase, function unknown: a transacetylase, function unknown

A regulatory gene A regulatory gene lacIlacI The LacI protein is a The LacI protein is a repressorrepressor of the of the lac lac operon operon

A promoter where RNA polymerase binds to transcribe A promoter where RNA polymerase binds to transcribe the operonthe operon

An operator site where the LacI repressor binds to An operator site where the LacI repressor binds to block transcriptionblock transcription

Arrangement of the Arrangement of the laclac operon operon

In the absence of lactoseIn the absence of lactose

In the absence of lactose, the repressor binds to In the absence of lactose, the repressor binds to operator site preventing binding of RNA operator site preventing binding of RNA polymerase and transcriptionpolymerase and transcription

In the presence of lactoseIn the presence of lactose The The laclac operon is operon is

an an inducible inducible operon operon

Lactose acts as Lactose acts as the inducerthe inducer

Lactose induces Lactose induces enzyme enzyme expression by expression by binding to the binding to the LacI repressor LacI repressor preventing its preventing its binding to the binding to the operator siteoperator site

The The argarg operon operon

A repressible operon: genes are A repressible operon: genes are transcribed until turned off or repressedtranscribed until turned off or repressed

Three structural genes Three structural genes argCBHargCBH encoding encoding enzymes in the arginine biosynthetic enzymes in the arginine biosynthetic pathwaypathway

In the absence of arginineIn the absence of arginine

The The arg arg operon is operon is transcribed transcribed and enzymes and enzymes for the for the synthesis of synthesis of arginine are arginine are producedproduced

In the presence of arginineIn the presence of arginine

Arginine acts as Arginine acts as a a corepressorcorepressor and represses and represses its own its own synthesissynthesis

Like feedback Like feedback inhibition but inhibition but acts on enzyme acts on enzyme synthesis rather synthesis rather than activitythan activity

Summary of gene regulationSummary of gene regulation

The production of many bacterial proteins The production of many bacterial proteins are controlled by regulating transcription are controlled by regulating transcription endproduct endproduct Functionally related genes are arranged in Functionally related genes are arranged in

operons to allow coordinate regulationoperons to allow coordinate regulation Operons can be Operons can be

Induced in the presence of substrateInduced in the presence of substrate Repressed in the presence of endproductRepressed in the presence of endproduct

Transcripition mechanisms can be more Transcripition mechanisms can be more complex with DNA sequences known as complex with DNA sequences known as activators and attenuators.activators and attenuators.