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Regulation
of
Enzyme
Synthesis |
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Regulation
of
Enzyme
Synthesis -
Bacteria
can
use
a
wide
range
of
substances
for
growth.
However,
the
ability
to
use
a
given
compound
depends
on
the
genetic
information
that
they
possess
which
is
necessary
to
synthesize
the
required
enzymes.
This
kind
of
regulation
of
enzyme
synthesis
was
first
described
by
J.
Monod
and
his
collaborators
in
1961
through
the
study
of
the
synthesis
of
the
enzymes
involved
in
the
utilization
of
lactose
by
E.coli.
This
bacterium
synthesises
high,
levels
of
these
enzymes
only
when
grown
in
a
medium
containing
lactose
as
a
carbon
source.
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The
enzymes
involved
are β-galactoside
permease,
(enzyme
involved
in
the
entry
of
lactose
into
the
cell)
f-galactosidase
and
transacetylase.
In
turn,
galactose
formed
by
the
action
of β-galactosidase
on
lactose,
induces
the
enzymes
responsible
for
the
metabolism
of
galactose.
Thus,
the
metabolism
of
one
primary
substrate
leads
to
the
induction
of
a
series
of
enzymes
that
are
necessary
for
the
complete
utilization
of
the
substrate.
In
this
case,
lactose
serves
both
as
an
inducer
as
well
as
a
carbon
source.
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There
are
other
inducers
which
are
not
metabolized
and
these
are
known
as
gratuitous
inducers.
Examples
of
such
inducers
for
enzymes
involved
in
lactose
degradation
in
E.Coli
are
isopropyl-β-thio-D
galactoside,
thiomethyl
galactoside
etc.
When
gratuitous
inducers
are
used,
enzyme
synthesis
continues
at
a
constant
rate
without
affecting
the
rate
of
growth
Any
compound
that
induces β galactosidase
synthesis
in
E.Coli
also
induces
the
synthesis
of
galactoside
permease
and
transacetylase
and
the
differential
rate
of
synthesis
of
these
enzymes
remains
constant.
This
type
of
synthesis
is
called
coordinate
induction.
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The
observations
by
Monad
and
others
that
enzyme
induction
and
repression
were
similar
but
opposite
phenomena
suggested
that
these
are
manifestations
of
a
fundamentally
similar
mechanism.
This
implied
that
the
precess
of
induction
and
repression
are
under
a
common
controlling
element
which
is
independent
of
the
structural
genes
that
code
for
the
different
enzymes.
A
genetic
study
of
a
variety
of
mutants
of
the
lactose
system
in
E.Coli
by
Jacob
and
Monad
led
to
the
identification
of
a
gene
that
regulate
the
rate
of
functioning
of
the
structural
genes
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It
was
proposed
that
the
product
of
this
regulatory
gene
(repressor)
combines
with
a
specific
region
of
the
DNA
called
the
operator
and
thus
regulates
the
rate
of
expression
of
the
structural
genes.
Support
to
this
hypothesis
came
from
the
study
of
mutants
in
the
operator
region
and
in
the
regulatory
gene.
For
example,
some
mutants
in
the
regulatory
gene
had
the
ability
to
synthesize
the
enzymes
in
the
absence
of
the
inducer.
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Similarly, some mutants in the operator region either produced no enzymes or produced excess (If the enzymes, whose level could not be further increased by the presence of the inducer. Genetic mapping of these mutations indicated that these are not linked. Further the observation that a single mutation in the operator region was found to affect the synthesis of three different enzymes involved in lactose metabolism led to the conclusion that it governs directly the rate of synthesis of the three enzymes by regulating the activity of three structural genes (genes that determine the proper sequence of amino acids in a peptide). This genetic unit of coordinate expression consisting of the operator and the structural genes was termed as the Operon.
According to this model of Jacob and Monod, the bacterial genome can be divided into a number of functional units of expression called the Operons which contain one or more structural genes. The transcription of these genes is under the control of a regulatory gene ,(R) and an Operator gene (O). The R gene is the structural gene for the synthesis of the repressor. Attachment of the repressor to the O gene was presumed to block transcription. In inducible systems, it was postulated that the inducer interacts with the repressor and brings about a steric modification resulting In the decreased affinity of the represor to the operator, which then allows transcription of the structural genes to occur at a higher rate.
Since the early description of this model based on studies with E.Coli and its lactose utilization system, there is a large amount of information that has accumulated regarding the regulation of enzyme synthesis which is mostly consistent with this classical model. The study of ß-galactosidase induction in E.Coli has provided much information about the nature of regulation of enzyme synthesis in bacteria. It is now generally agreed that induction of enzyme, synthesis is mediated by non catalytic allosteric proteins called repressors. These proteins are products of specific regulatory genes and they control enzyme synthesis in a negative manner by binding to the bacterial DNA at a site near the structural genes and thus prevent their transcription. The protein nature of the repressors has been confirmed and at least five or six different repressors have been now purified. Among these, the ß-galactosidase repressor is the. best characterized and consists of a protein of four identical subunits each with a molecular weight of 37,000 and with 347 amino acids. It is now known that these repressors bind to a specific nucleotide sequence of the operator region. The nucleotide sequence of the lac operator region where the repressor binds has been found to consist of 21 base pairs with 16 bases related by a two fold axis of symmetry centred at the 11th base pair . Such symmetry has been found to allow binding of two subunits of the tetrameric lac repressor simultaneously to the operator.
Variations in the amount of protein in a cell generally reflects the rates of synthesis which in turn relates to the number of m-RNA templates available for translation. It has been found that the number of m-RNA templates in cells synthesisil1g enzymes is much larger than in those which do not. For example, during maximal ß-galactosidase synthesis, the cells contain 35-50 ß-galactosidase m-RNA molecules while in the absence of the inducer, the cell may contain one or less number of molecules.
Repressor molecules exist in both an active and an inactive form depending on whether they are associated with the inducers. In some cases, as in the lac repressor, attachment of in inducer in activates the repressor while in an another case, as with the arabinose repressor, activation of the repressor leads to enzyme synthesis. Also, the addition of amino acids to cells activates repressors which control the synthesis of the enzymes involved in the biosynthesis of amino acids. This immediately shuts off synthesis of specific m-RNA molecules.
In the interaction of the repressors with corepressors (metabolites which by their combination with repressors specifically inhibit enzyme synthesis) or inducers, no covalent bond formation occurs and therefore the weak bonds are rapidly made and broken allowing the repressors to be either active or inactive depending on the physiological need.
The lactose operon is a classical example of an operon under negative control i.e. the product of the regulatory gene controls the operator functions negatively by not allowing transcription. Induction of enzymes may also involve a type of control by the allosteric protein, which has both a positive and negative component . For example, E.coli can metabolise arabinose as the carbon source by the induced synthesis of four enzymes. The arabinose repressor is essential for the functioning of the arabinose operon.
In the presence of arabinose which is the inducer, the repressor molecule binds to the operator and allows transcription while in the absence of arabinose, the repressor blocks transcription like the lactose repressor When arabinose binds to the repressor, the latter undergoes a conformational change and is converted into an activator and thereby triggers transcription. This type of control is termed as positive control. In bacteria, both positive and negative type of control mechanisms are now known to operate in regulating macromolecular synthesis.
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