MIC 428 Lecture

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MIC 428 - Lecture 14 Regulation of Gene
Expression (1st part)
Requires review of Chapter 7 (Brocks book)
before coming to class.
Outline

• The complexity of regulation.
• Overview of regulation.
• Constitutive and inducible enzymes.
• Major modes of regulation in the cell
• Control of enzyme activity. Post translational
  control.
• Feedback inhibition.
• Allosteric regulation and rate-limiting enzymes.
• Types of allosteric regulation.
• Isoenzymes.

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Outline continuation
Covalent modification and the case of glutamine
synthase. 2. Control of the amount of enzyme
present transcriptional control and
translational control. DNA binding proteins. The
importance of protein-nucleic acid
interactions. Specific and non-specific
interactions Histones and sequence-specific
proteins. Protein domains and interaction with
inverted repeats. Structure of DNA binding
proteins Helix-turn-helix, Zn finger, Leucine
zipper. Negative control of transcription
repression and induction.
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Complexity of regulation
Hundreds of different enzymatic reactions
happening simultaneously.
Need to respond rapidly to changes in their
environment.
Complicated developmental pathways.
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Some enzymes are needed in the same amount under
any growth condition.
Some enzymes are not permanently needed in the
same amount under any growth condition.
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Two major modes of regulation
Control of enzyme activity
After the enzyme has been produced
Post translational control
Control of the amount of an enzyme
At the level of transcription
At the level of translation
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Overview of regulation
The product of gene A is enzyme A, which is
synthesized constitutively and carries out its
reaction. Enzyme B is also synthesized
constitutively but its activity can be inhibited.
The synthesis of the product of gene C can be
prevented by control at the level of translation.
The synthesis of the product of gene D can be
prevented by control at the level of
transcription.
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Regulation of enzyme activity
Inhibiting enzyme activity
Enzymes are synthesized with full enzymatic
activity and then the activity is reduced or
inhibited by certain compounds in the cell.
We have seen some of these cases. Do you remember
any?
These compounds are usually related to the
metabolic pathway in which the enzyme functions.
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The EM pathway
Inhibited by ATP Inhibited by high levels of
glucose-6-P Inhibited by Ala, an intermediate
synthesis product from pyruvate.
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Noncovalent Enzyme Inhibition
Feedback inhibition
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Feedback inhibition of enzyme activity
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How is it possible for the end product of a
pathway to inhibit the enzyme that acts on a
substrate quite unrelated to it?
By means of allosteric regulation
The regulated enzyme is referred to as the
rate-limiting enzyme.
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Mechanism of enzyme inhibition by an allosteric
effector
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In branched metabolic sequences, the beginning of
each branch is often controlled allosterically
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Feedback inhibition in a branched biosynthetic
pathway
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Types of allosteric regulation
1. Simple feedback inhibition. 2. Concerted
feedback inhibition. 3. Sequential feedback
inhibition.
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1. Simple feedback inhibition.
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2. Concerted feedback inhibition.
A
B
C
D
E
F
More than one metabolite interacts with the enzyme
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3. Sequential feedback inhibition.
A
Two or more metabolites interact with an enzyme
B
C
F
D
G
E
H
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Some biosynthetic pathways are regulated by
isoenzymes that catalyze the same reaction but
are subject to different regulatory controls.
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The common pathway leading to the synthesis of
the aromatic amino acids contains three isozymes.
Each of these enzymes is specifically
feedback-inhibited by one of the aromatic amino
acids. Note how an excess of all three amino
acids is required to completely shut off the
synthesis of DAHP.
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Covalent modification of enzymes glutamine
synthetase (GS)
Enzyme activity highly controlled
GS activity is controlled by concerted feedback
inhibition by nine different compounds including
amino acids and compounds involved in nucleotide
metabolism.
GS activity independently regulated by covalent
inhibition by glutamine and ketoglutarate.
Complex molecule with 12 subunits.
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When we consider posttranslational modification
of proteins
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Processing of preproinsulin
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Protein splicing
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Protein-nucleic acid interactions
Please read section 8.4 from the book since I
wont cover the topic in detail but you will be
tested on it.

• Protein-nucleic acid interactions are central to
• Replication
• Transcription
• Translation
• Regulation of these processes.

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Protein-nucleic acid interactions

• Two types of interactions
• Non-specific
• Specific.

Example of non-specific interaction histones.
Example of specific interaction
sequence-specific proteins.
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Sequence-specific proteins
To achieve specificity the protein must interact
simultaneously with more than one nucleic acid
base.
Do you remember what inverted repeats are?
Frequently DNA binding proteins combine
specifically with inverted repeats in DNA.
Many such proteins are dimers that combine
specifically with two sites on the DNA.
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Sequence-specific DNA-binding proteins
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Structure of DNA binding proteins

• Helix-turn-helix
• Zinc finger
• Leucine zipper

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The helix-turn-helix structure of some DNA
binding proteins
(a) A simple model of the helix-turn-helix
elements.
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(a) The zinc finger structure. The amino acids
holding the Zn2 ion always include at least two
cysteine residues (C) with the other residues
being histidine (H). (b) The leucine zipper
structure. The leucine residues (shown in yellow)
are always spaced exactly every seven amino
acids. The interaction of the leucine side chains
helps hold the two helices together.
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Negative control of transcription
If one gene is transcribed more frequently
compared to another one, there will be a greater
abundance of mRNA and a greater abundance of the
final protein.
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Do you remember what consensus sequences are? And
Shine-Dalgarno sequences? If not, review Chapter
7.
High levels of transcription can result because a
gene has a promoter very close to the consensus
sequence and high levels of translation can
result if the genes Shine-Dalgarno has extensive
complementarity with 16S rRNA.
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But we will discuss how the level of
transcription of an individual gene can be
changed in a regulated manner.
Enzyme repression
Often the enzymes catalyzing the synthesis of a
specific product are not synthesized if this
product is present in the medium. Ej enzymes
involved in the synthesis of Arginine.
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Repression of enzymes involved in arginine
synthesis by addition of arginine to the medium
Note that the rate of total protein synthesis
remains unchanged.
Specific effect
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Enzyme repression is a very widespread phenomenon
in bacteria
In almost all cases the final product of a
particular biosynthetic pathway represses the
enzymes of that pathway.
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Enzyme induction
It is the synthesis of an enzyme only when the
substrate is present.
ß- galactosidase
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Induction of the enzyme b-galactosidase on the
addition of lactose to the medium
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Enzymes involved in the catabolism of carbon and
energy sources are often inducible.
The substance that initiates enzyme induction is
the inducer. The substance that represses
production is called a corepressor. Collectively
called effectors.
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Analogs of these substances may induce or repress
even though they are not substrates of the enzyme.
IPTG (Isopropylthiogalactoside)
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