Download Biol115_2014_Lecture 10_Prokaryotic Gene Regulation

Biol115
The Thread of Life"
Lecture 10"
Regulation of gene expression I:
prokaryotic systems"
Cheese is milk's leap toward immortality"
~Cliff Fadiman!
Objectives"
•  Describe the organization of bacterial DNA into operons.!
•  Compare and contrast the operations of a repressible and an
inducible operon."
•  Key terms: allosteric effector, allosteric protein, inducible
operon, operator, operon, promoter, repressible operon,
repressor"
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Principles of Biology"
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Chapter 18 Prokaryotic Gene regulation"
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Overview: conducting the genetic
orchestra"
•  The genome contains all of the information needed for the
growth and development of individuals."
•  Gene expression refers to the use of DNA sequences to
synthesise RNA and proteins."
•  Some genes are expressed all the time (constitutive or
housekeeping), while others are expressed only when needed
(regulated)."
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Gene expression in prokaryotes
Since bacteria have no nucleus, transcription and translation can occur in the
same space.
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Bacteria often respond to environmental
change by regulating transcription"
•  Natural selection has favoured bacteria that produce only the
products needed by that cell"
•  A cell can regulate the production of enzymes by feedback
inhibition or by gene regulation"
•  Gene expression in bacteria is controlled by the operon model"
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Tryptophan synthesis
A diagram shows that tryptophan synthesis is
inhibited by binding of tryptophan to an enzyme
required for its synthesis. In the absence of
tryptophan, E. coli activates a metabolic pathway to
begin tryptophan synthesis from a precursor. When
tryptophan is abundant, either from the environment
or through synthesis, the metabolic pathway
undergoes feedback inhibition. Tryptophan inhibits
the activity of an enzyme in its synthesis pathway.
Additionally, tryptophan can repress the tryptophan
synthesis pathway by negatively regulating the gene
expression of the enzymes necessary for tryptophan
synthesis. 5 enzymes are required for tryptophan
synthesis. Tryptophan inhibits both an enzyme
in the pathway and the expression of the
genes encoding the enzymes.
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Operons: The basic concept"
•  A cluster of functionally related genes can be under coordinated
control by a single on-off “switch”"
•  An operon is the entire stretch of DNA that includes the
operator, the promoter, and the genes that they control"
•  The regulatory “switch” is a segment of DNA called an operator
usually positioned within the promoter"
•  The operon can be switched off by a protein repressor!
•  The repressor prevents gene transcription by binding to the
operator and blocking RNA polymerase"
"
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Operons: The basic concept"
•  The operon can be switched off by a protein
repressor!
•  The repressor prevents gene transcription by
binding to the operator and blocking RNA
polymerase"
•  The repressor is the product of a separate
regulatory gene!
•  The repressor can be in an active or inactive form,
depending on the presence of other molecules"
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http://tinyurl.com/kzpscbe"
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Contents of an operon"
"Promoter: RNA polymerase binding site"
"Repressor: regulatory protein that binds DNA (see operator)"
"Operator (controlling site): DNA region that is downstream of
promoter and to which the repressor binds"
"Coding sequences: also known as structural genes, which
specify the expression of proteins involved in a specific metabolic
pathway"
"Terminator: DNA sequence which signifies the end of an operon."
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More definitions"
•  Regulator gene – A gene that
codes for a product (typically
protein) that controls the
expression of other genes (usually
at the level of transcription)."
•  Structural gene – A gene that
codes for any RNA or protein
product other than a regulator."
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The concept of operons"
http://www.nature.com/scitable/content/33138/pierce_16_3_FULL.jpg"
Note: for this lecture, assume that the regulator protein = repressor"
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The repressor protein – a shape shifter!"
•  Each operon is controlled by its own repressor protein."
•  Repressor proteins are called allosteric proteins. Allosteric
proteins are proteins that change their shape when bound to a
specific molecule called an allosteric effector."
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The trp (tryptophan) operon"
•  If amino acids are present in the growth medium E. coli will “import” amino
acids before it makes them."
•  By default the trp operon is on and the genes for tryptophan synthesis are
transcribed"
•  When tryptophan is present, it binds to the trp repressor protein, which
causes it to change its shape and thus turn off the operon. "
•  The repressor is active only in the presence of its corepressor,
tryptophan; thus the trp operon is turned off (repressed) if tryptophan
levels are high"
•  A corepressor is a molecule that cooperates with a repressor protein to
switch an operon off"
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Insert Fig. 18-3 P 355"
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Trp operon – genetic terminology"
Element!
Function!
Notes!
trpA-E"
Genes that encodes enzymes involved in
tryptophan synthesis"
trpR"
Not a part of the trp
Gene that encodes the trp repressor protein
operon, but regulates
(when bound to DNA (operator), this proteins
transcription of the trp
prevents transcription at the trpoperon)"
operon"
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Structural genes of the
trp operon"
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Lactose metabolism"
•  Lactose = major sugar of milk "
•  Lactose is an uncommon
nutrient for bacteria"
•  Enzymes for lactose
metabolism are normally not
expressed (turned off)."
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Lactose"
•  A disaccharide of glucose and
galactose"
• 3 enzymes are involved:"
"
"
1. permease (lacY)- "
allows lactose to enter bacterial cells"
2. β-galactosidase (lacZ) - breaks
down lactose to glucose and galactose"
3. transacetylase (lacA) - unknown
function but assists lactose breakdown"
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The lac operon
Consists of a promoter (P), operator (O) and three structural genes (lacZ, lacY and lacA). A
separate gene, lacI, encodes the Lac repressor.
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The lac operon
In the absence of lactose, the Lac repressor is bound to the operator, preventing RNA
polymerase from transcribing the structural genes.
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The lac operon
In the presence of lactose, some allolactose is produced. Allolactose binds to allosteric site of
Lac repressor, changing its conformation so that it can no longer bind the operator. This allows
RNA polymerase to transcribe the structural genes.
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Repressor does not completely
inhibit transcription!
So very small amounts of the
enzymes are made!
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Lac repressor protein (violet) forms a
tetramer which binds to DNA, causing a
loop to form in the DNA. As a result
expression of the lac operon is turned off. "
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Lac operon – genetic terminology!
Element!
lacZ"
Function!
Gene that encodes the β-galactosidase
protein (enzyme that breaks down lactose to
glucose and galactose)"
Notes!
lacY"
Gene that encodes the permease protein (a Structural genes of the
membrane protein that imports lactose into
lac operon"
bacterial cells)"
lacA"
Gene that encodes the transacetylase
protein (unknown function)"
lacI"
Gene that encodes the lac repressor
protein (when bound to DNA (operator), this
proteins prevents transcription at the lac
operon)"
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Not a part of the lac
operon, but regulates
transcription of the lac
operon"
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Operon!
Conditions!
Repressor!
Low intracellular Is inactive and does not bind to
tryptophan"
operator sequence on DNA"
trp!
Result!
RNA polymerase transcribes trp
operon genes, which results in
tryptophan biosynthesis"
High
intracellular
tryptophan"
Is active. Tryptophan
(corepressor) binds to repressor
and induces the repressor
protein to bind to operator
sequence on DNA."
No extracellular
lactose"
RNA is obstructed from
Is active. It binds to the operator transcribing lac operon genes.
sequence on DNA."
No lactose metabolising
enzymes are synthesised."
High
extracellular
lactose"
Is inactive. Allolactose (inducer)
binds to repressor and reduces
its affinity for DNA. Repressor
protein falls off DNA."
lac!
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RNA polymerase is obstructed
from transcribing trp operon
genes. Tryptophan
biosynthesis stops."
RNA polymerase is free to
transcribe lac operon genes.
Concentration of lactose
metabolising enzymes
increases."
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Repressible and inducible operons"
•  A repressible operon is one that is usually on; binding of a
repressor to the operator shuts off transcription"
•  The trp operon is a repressible operon.!
•  An inducible operon is one that is usually off; a molecule
called an inducer inactivates the repressor and turns on
transcription"
•  The lac operon is an example of an inducible operon."
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Repressible and inducible operons"
•  Inducible enzymes usually function in catabolic pathways; their
synthesis is induced by a chemical signal"
•  Repressible enzymes usually function in anabolic pathways;
their synthesis is repressed by high levels of the end product"
•  Regulation of the trp and lac operons involves negative control
of genes because operons are switched off by the active form of
the repressor"
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Summary"
•  In inducible regulation, the gene is regulated by the presence
of its substrate (the inducer) – e.g. allolactose and the lactose
operon."
•  In repressible regulation, the gene is regulated by the product
of its enzyme pathway (the corepressor) – e.g. tryptophan
metabolism."
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You should now be able to:"
1.  Explain the concept of an operon and the function of the
operator, repressor, and co-repressor."
2.  Explain the adaptive advantage of grouping bacterial genes
into an operon."
3.  Explain how repressible and inducible operons (using the trp
and lac operons as examples) differ and how those
differences reflect differences in the pathways they control."
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