Microbial Genetics

1
Microbial Genetics

• The Operon
• Regulation of bacterial gene expression

2
Controlling enzymes

• We know that
• Bacterial cells carry out a huge number of
  chemical reactions catalyzed by enzymes
• And .. Bacterial cells respond rapidly to
  changing environments e.g., presence of a new
  carbon source
• So .
• control of Enzymes is VERY important and must be
  efficient

3
Enzyme Regulation

• Enzyme activity can be regulated by feedback
• inhibition of synthesized enzyme
• BUT making enzyme requires energy a a bacterium
  cannot make ALL enzymes ALLTHE TIME
• SO it is better to stop synthesizing enzymes that
  are not needed
• THUS, many genes such as those for enzymes
  involved in lactose metabolism or amino acid
  production are regulated and only expressed when
  required

However, 60-80 of genes are not regulated and
are termed Constitutive
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Regulation of gene expression
Constitutive Enzymes No regulation
Enzyme Induced Enzyme Repressed
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Regulation of gene expression

• The process of transcription plays an important
  role in the ability of bacteria to respond to
  changing environments mRNA compared to DNA is
  inherently unstable.
• Thus, the expression levels of certain proteins
  can be controlled at the level of transcription
  (mRNA synthesis).

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Transcriptional control

• Two genetic control mechanisms which regulate
  transcription of mRNA and, therefore the
  synthesis of corresponding protein
• Induction-turning on gene expression
• Repression-turning off gene expression

7
Repression

• Inhibition of gene expression
• Often in response to an excess of endproduct,
  shuts down synthesis of the enzyme
• Mediated by regulatory proteins called repressors
• Block RNA polymerase from initiating
  transcription
• Repressible genes are transcribed until they are
  repressed

8
Induction

• Turns on transcription of a gene
• A substance which acts to induce transcription of
  a gene is an inducer, often the substrate
• Enzymes synthesized in the presence of inducers
  are called inducible enzymes
• Example lactose utilization genes
• Inducible genes are not transcribed until they
  are induced

9
The operon

• Multiple genes are arranged in the same
  orientation and are closely linked on the DNA
• Genes in an operon are transcribed on a single
  RNA transcript, but are translated individually
  to form multiple proteins
• A mechanism for coordinate control of genes
  involved in a single process by regulating
  transcription of the operon mRNA

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The operon (cont)

• The term operon not only includes the
  structural genes in the operon but also the
  regulatory sequences controlling transcription
• Promoter-site of RNA polymerase binding
• Operator-site of binding of a regulatory protein

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Operons and regulation

• About 27 of E. coli genes are in operons
• Many of the genes arranged in operons are
  regulated
• For example,
• genes for the lactose utilization are controlled
  by the presence of lactose
• genes for pathogenesis are often controlled by
  temperature or physiologic factor

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Two regulated operons

• Two E. coli operons reflect how operons in
  general are regulated
• The lac operon
• Encodes inducible catalytic enzymes involved in
  the lactose utilization and uptake
• The arg operon
• Encodes repressible anabolic enzymes involved in
  the production of the amino acid arginine

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The lac operon

• Three structural genes
• lacZ ß-galactosidase splits lactose to glucose
  and galactose
• lacY a permease involved in transport of lactose
• lacA a transacetylase, function unknown
• A regulatory gene lacI
• The LacI protein is a repressor of the lac operon
  
• A promoter where RNA polymerase binds to
  transcribe the operon
• An operator site where the LacI repressor binds
  to block transcription

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Arrangement of the lac operon
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In the absence of lactose

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

16
In the presence of lactose

• The lac operon is an inducible operon
• Lactose acts as the inducer
• Lactose induces enzyme expression by binding to
  the LacI repressor preventing its binding to the
  operator site

17
The arg operon

• A repressible operon genes are transcribed until
  turned off or repressed
• Three structural genes argCBH encoding enzymes in
  the arginine biosynthetic pathway

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In the absence of arginine

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

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In the presence of arginine

• Arginine acts as a corepressor and represses its
  own synthesis
• Like feedback inhibition but acts on enzyme
  synthesis rather than activity

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Summary of gene regulation

• The production of many bacterial proteins are
  controlled by regulating transcription endproduct
  
• Functionally related genes are arranged in
  operons to allow coordinate regulation
• Operons can be
• Induced in the presence of substrate
• Repressed in the presence of endproduct
• Transcripition mechanisms can be more complex
  with DNA sequences known as activators and
  attenuators.