Download THE LAC OPERON

THE LAC OPERON
BEGINS
Hello there boys and girls. I’m Mr. Milk and I’m here to present the
Lac Operon. The Lac Operon is found in bacteria and basically acts
like and on –off switch for the bacteria genes. Operons are special
because normally, each gene has its own promoter. Here several
genes required for function, but only have the one promoter
Do you see the
agar plates with
the bacteria
growing on
them? Those
bacteria will only
make the enzyme
to break down
lactose is lactose
is added
This bacteria is on a
medium with minimal
nutrients and displays
limited growth. The
bacteria are not producing
lactose digesting enzymes.
If the same petri dish is flooded
with lactose, the bacteria will
turn on the gene to make the
enzyme to digest lactose. They
grow and divide to fill the
entire dish.
Note: lactose cannot be used for cellular metabolism unless it
is broken into glucose and galactose – glucose into glycolysis
There are names for particular agar broth mixtures that
are used to test for bacteria with specific functional
genes and thus proteins. It is one of the mechanisms of
bacteria identification in lab cultures. MacConkey lactose
and tetrazolium lactose media both both contain lactose
and bacteria that grow on these plates have to have the
enzyme β-galactosidase. That includes lac Z and Y.
This is the process of creating streak
plates of bacteria. The agar may
contain various nutrients that can be
used to identify if the bacteria will
grow on the media. If colonies of the
sample are seen growing, they can use
the media. If no sample colonies are
seen, the bacteria cannot utilize the
nutrient resource.
If you put some of those bacteria into my milk
container, they will also turn on the gene to
produce the lactose digesting enzymes. How do
the bacteria know to turn on that gene? They
don’t have brains, yet it seems that they are
making a decision on which enzymes to
manufacture inside their cells.
The bacteria utilize
something called an
operon to control gene
expression. They can turn
on or off the lactose
digesting gene with the
LAC OPERON. Let me
explain how it works
Jacob and Monad (France)conducted this
experiment with E coli bacteria . They
researched the Lac operon model we will
study. They proposed that genes with
related functions are grouped together as
units called operons. There are others
such as the tryptophan operon etc.
Hope you still
remember milk
contains lactose!
In bacteria, as well as other
organisms, this is called
REGULATION of gene expression.
Genes that produce specific proteins
must turn on or off at the right times
and in the right places. The Lac
operon is a study in bacteria (a
simple system) used to improve our
understanding of how this occurs.
Eukaryotic Gene Regulation is very
complicated and will be covered in
another presentation.
First we must start with the DNA. DNA is not just pure DNA
on the chromosome. Rather it is associated with vast
amounts of protein which is involved in control of the genes
on the DNA. In our model of the Lac Operon , we will
represent some of the associations with DNA and protein by
naming the DNA regions.
In order for DNA to be transcribed, it must have a region
(called the promoter) for the RNA polymerase to join.
Promoter
RNA
p
DNA CODE -3’5’
RNA
polymerase
RNAp
Transcribed to mRNA ex 5’AUGGGCGGAAAUUUGGA3’
Translated to amino acids in sequence to make a protein
One of the most important aspects of the
operon is the control system. The lac operon
utilizes a repressor protein that stops the
transcription of the lac operon by blocking the
gene access by RNA polymerase. We’ll see how
this is done later.
In the process below the repressor protein is made
Promoter
RNA
p
DNA CODE -3’5’
RNA
polymerase
RNAp
Transcribed to mRNA ex 5’AUGGGCGGAAAUUUGGA3’
Translated to amino acids in sequence to make repressor protein –
simulated molecular shape below
Repressor
protein
Note there is a codon but no tRNA
for the stop signal.
The repressor protein has a specific shape
that bonds with a part of the DNA on the Lac
operon called the OPERATOR. The operator is
locate d in between the PROMOTER and the
STRUCTURAL gene of the lactose breakdown
genes.
Promoter
RNA
p
Operator
Repressor
protein
Lactose breakdown enzyme genes
Hey, I’m blocked. I
can’t engage the
DNA to begin any
copy process!
The lactose enzyme
information is not
transcribed. No lactose
breakdown enzymes
are made. This gene is
effectively turned OFF
The lac operon is an INDUCIBLE Operon.
Addition of an inducer(signal molecule)
turns the gene ON. The inducer is the
chemical that needs to be digested. In this
case lactose is the inducer.
The inducer bonds to the repressor!
Promoter
Operator
Lactose breakdown enzyme genes
Transcribed to mRNA ex 5’AUG..3’is called polycistronic –many genes 1 mRNA
Translated to amino acids in sequence to make LACOSE DIGEST ENZ
RNA
p
Lactose
Repressor
protein
Hey, I’m blocked. I
can’t engage the
DNA to block RNA
polymerase!
The lactose enzyme
information is transcribed.
Lactose breakdown enzymes
are made. This gene is
effectively turned ON
There are actually three enzyme breakdown genes of the Lac
operon. They are shown below in their 3-D model structures.
Each will participate in the breakdown of lactose into the two
simple sugars glucose and galactose
Remember, lactose is
only useful to the
bacteria if they can
break it down and put
the sugars into
glycolysis. Glucose
enters easily.
Galactose is just an
isomer of glucose and
can be converted.
Isomers again.
See I told you it
would be
important!
Coded for by Lac Z
part of operon
Coded for by Lac A part of
operon.Bacteria are
capable of catabolism of
lactose even without this
enzyme
Coded for by Lac Y
part of operon
LACTOSE
ENZYMES
GLUCOSE
GALACTOSE
Now once the enzymes are made by the process of transcription
and translation, the enzymes start doing their job breaking
down the lactose. They continue to do this until all the
lactose is used up. When this happens the bacteria then
shut off the gene again.
NO MORE LACTOSE
RNA
p
Promoter
Operator
Repressor
protein
NO MORE NEED FOR ENZYMES
NO MORE INDUCER TO
BLOCK REPRESSOR
Lactose breakdown enzyme genes
Hey, I’m blocked
again. I can’t
engage the DNA to
begin any copy
process!
There is no need to
make lactose
breakdown enzymes
anymore. No more
lactose is left. This gene
is effectively turned
OFF
And that’s the story of how an
INDUCIBLE OPERON works. Here is
an example using different shape
representation
The repressor gene is technically
not part of the operon and may
be physically separated by
distance from the operon.
This region is all part of the Lac operon which
codes for three protein enzymes.
Above is another diagram of the lac operon.
Once the bacteria are exposed to different sugars, their
operons have to either produce enzymes or not. Some
control of the Lac operon is due to cAMP –cyclic
Adenosine Monophosphate. cAMP is made when
glucose levels are low and it acts to TURN ON or
ACTIVATE the Lac operon. The Lac operon, however, will
still not turn on unless lactose is present. In this case two
conditions must be met to turn on the operon.
Cyclic AMP is found in both prokaryotes and eukaryotes. It is
derived from ATP but is not used for energy but rather as a
secondary messenger to control cell activities.(signal transduction)
CAP is
catabolite
activator
protein and
will greatly
increase the
production of
galactosidase
in the
absence of
glucose. It
needs cAMP
to activate.
Lac operon turned ON when glucose
low (high cAMP) and lactose present
Some operons work differently than the
Lac operon. These are called Repressible
Operons. The example is the Trp OPERON
In the Trp operon, tryptophan is synthesized because the
gene is always turned on unless the inducer is present.
Normally, the gene is turned off because tryptophan is
present and this bonds to the repressor (tryptophanrepressor complex) changing its shape ( an allosteric
change) and causing it to bind to the operator(active
repressor). It is a repressible operon because if typtophan
is not present, the gene will be on. The repressor is not
the right shape to bond to the operator unless the
co-repressor(trp) is present
REPRESSIBLE - NO INDUCER – STRUCTURAL GENE IS TURNED ON!!
Opposite to inducible operon –
The trp operon a repressible operon because it is
always ON, but it can be turned OFF.
Trp operon includes 5 genes –make 5 enzymes that
convert molecules into tryptophan
Bacteria need tryptophan to make cell proteins
( E. Coli) and needs a constant supply.
Tryptophan making genes are always turned on by
the operon, but making typtophan takes energy
If the bacteria find tryptophan in the environment,
why would they use energy making something that is
readily available? They shut the operon off!
This is accomplished by having the repressor only
activated into the matching shape of the operator if
tryptophan is present. This shuts the genes off!
So the tryp operon is different because it is always on unless
tryptophan can be found in the environment. If tryptophan is
available, it bonds to the repressor making the repressor have the
correct shape to bond to the operator and block the transcription of
the trp genes. This is shown by the diagram below.
Trp is called a
corepressor! It
is necessary for
it to bond to the
repressor for
the repressor to
engage the
operator. Trp is
not an inducer!
Lets try some questions. In what
organisms do you find operons?
THINK THINK THINK
E. Coli is a bacteria
Operons are located in bacteria genomes
Regarding the trp
operon; w hen
tryptophan is not
present, normal
transcription
continues. Is it an
example of
negative or
positive
feedback?
All prokarytotes contain operons
Operons are only found in the prokaryotes
THINK THINK THINK
Trp operon is a repressible operon
When trp is present genes turn off transcription
When trp is absent genes turn on transcription
Negative feedback means what is produced
accumulates and shuts down the system.
This is an example of negative feedback
One last thing. Lets take a fast look at the experiment that starting
this whole idea of the operon. Remember the French Scientists Jacob
and Monod were the ones responsible.
Monad in the 1940’s noticed that if glucose and lactose
were both provided, glucose would be metabolized first
(growth phase I, and then lactose (growth phase II).
This phenomenon is called diauxis
First Jacob and Monod grew the bacteria on a culture media containing
radioactive amino acids.
Then Jacob and Monod added lactose to a culture of E.coli cells, and found
that lactose was metabolized. Radioactive amino acids were incorporated
into the enzyme beta-galactosidase.
This was because the inducer(the lactose) removes the repressor and allows
the transcription and translation of the beta-galactosidase gene.
The product, β-galactosidase then starts to digest lactose into galactose and
glucose.
Bye for now.
Lac Operon
END