Lac Operon

S U G I YAT I N I N G R U M
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 Lac Operon

 The lac operon (lactose operon) is an operon required for

the transport and metabolism of lactoses in E.coli and
many other enteric bacteria.
 Although glucose is the preferred carbon source for most
bacteria, the lac operon allows for the effective digestion of
lactose when glucose is not available through the activity of
beta-galactosidase.
 The lac operon of E. coli contains genes involved in lactose
metabolism. It's expressed only when lactose is present
and glucose is absent.
 Structure of Lac Operon

The lac operon consists of :

1. Three structural gene : Lac Z, Lac Y, Lac A
2. Operator : Negative regulatory site bound by the lac repressor protein. The
operator overlaps with the promoter, and when the lacrepressor is bound, RNA
polymerase cannot bind to the promoter and start transcription.
3. Promotor : the binding site for RNA polymerase
4. CAP Binding Site : is a positive regulatory site that is bound by Catabolite Activator
Protein (CAP). When CAP is bound to this site, it promotes transcription by helping
RNA polymerase bind to the promoter.
5. Lac I : an allosterically regulated repressor
 3 structural gene

The three structural genes are: lacZ, lacY, and lacA

 lacZ encodes β-galactosidase (LacZ), cleaves the disaccharide lactoses into glucose and galactose.
 lacY encodes β-galactoside permease (LacY), to transport lactose into the cell using a proton gradient in
the same direction.
 lacA encodes β-galactoside transacetylase (LacA), an enzyme that transfers an acetyl group from acetyl-
CoA to β-galactosides
 What makes the lac operon turn on?

 E. coli bacteria can break down lactose, but it's not their
favorite carbon source. If glucose is around, they would
much rather use that. Glucose requires fewer steps and
less energy to break down than lactose. However, if
lactose is the only sugar available, the E. coli will go right
ahead and use it as an energy source.
 To use lactose, the bacteria must express the lac operon
genes, which encode key enzymes for lactose uptake
and metabolism. To be as efficient as possible, E. coli
should express the lac operon only when two conditions
are met:
Lactose is available, and Glucose is not available
When does the lac operon really turn on?

The lac operon will be expressed at high levels if two

conditions are met :

 Glucose must be unavailable: When glucose is unavailable, cAMP

binds to CAP, making CAP able to bind DNA. Bound CAP helps RNA
polymerase attach to the lac operon promoter.
 Lactose must be available: If lactose is available, the lac repressor will
be released from the operator (by binding of allolactose). This allows
RNA polymerase to move forward on the DNA and transcribe the
operon.
 These two events in combination – the binding of the activator and the
release of the repressor – allow RNA polymerase to bind strongly to the
promoter and give it a clear path for transcription. They lead to strong
transcription of the lac operon and production of enzymes needed for
lactose utilization.
What will happen if lactose exists and doesn’t exist?

 The lac repressor is a protein that

represses (inhibits) transcription of
the lac operon.
 When lactose is not available,
the lac repressor binds tightly to
the operator, preventing
transcription by RNA polymerase.
 When lactose is available, some
molecules will be converted to
allolactose inside the cell,
the lac repressor loses its ability to
bind DNA because Allolactose
binds to the lac repressor and
makes it change shape so it can no
longer bind DNA.
What will happen if glucose exists and doesn’t
exist?

 CAP isn't always active (able to bind

DNA). Instead, it's regulated by a
small molecule called cyclic
AMP (cAMP).
 cAMP is a "hunger signal" made
by E. coli when glucose levels are
low. cAMP binds to CAP, changing its
shape and making it able to bind DNA
and promote transcription.
 Without cAMP, CAP cannot bind DNA
and is inactive. CAP is only active
when glucose levels are low (cAMP
levels are high). Thus, the lac operon
can only be transcribed at high levels
when glucose is absent.
what causes low glucose concentration, high cAMP?

• The operon is inducible by lactose

to the maximal levels when cAMP
and CAP form a complex.
a) when glucose concentrations are
low, an enzyme called adenylate
cyclase is active and able to
convert ATP into cAMP
b) when glucose concentrations are
high, an enzyme called adenylate
cyclase is nonactive and cannot
convert ATP into cAMP
Activation Adenylate Cyclase
 4 Possible situation

 Situation 1

Glucose present, lactose absent: No transcription of the lac operon

occurs. That's because the lac repressor remains bound to the operator
and prevents transcription by RNA polymerase. Also, cAMP levels are low
because glucose levels are high, so CAP is inactive and cannot bind DNA.
 4 Possible situation

 Situation 2

Glucose present, lactose present: Low-level transcription of the lac operon

occurs. The lac repressor is released from the operator because the inducer
(allolactose) is present. cAMP levels, however, are low because glucose is
present. Thus, CAP remains inactive and cannot bind to DNA, so
transcription only occurs at a low, leaky level.
 4 Possible situation

 Situation 3

Glucose absent, lactose absent: No transcription of the lac operon occurs.

cAMP levels are high because glucose levels are low, so CAP is active and
will be bound to the DNA. However, the lac repressor will also be bound to the
operator (due to the absence of allolactose), acting as a roadblock to RNA
polymerase and preventing transcription.
 4 Possible situation

 Situation 4

Glucose absent, lactose present: Strong transcription of the lac operon

occurs. The lac repressor is released from the operator because the inducer
(allolactose) is present. cAMP levels are high because glucose is absent, so
CAP is active and bound to the DNA. CAP helps RNA polymerase bind to the
promoter, permitting high levels of transcription.
Summary of lac operon responses
 References

 Berg, J. M., Tymoczko, J. L., and Stryer, L. (2002). Prokaryotic DNA-binding proteins bind
specifically to regulatory sites in operons. In Biochemistry (5th ed., section 31.1). New York,
NY: W. H. Freeman. Retrieved http://www.ncbi.nlm.nih.gov/books/NBK22512/.
 Bergmann, D. C. (2011). Gene regulation: How to only make RNA (and protein) at the right time
and place. In BIO41: 2011 Molecular biology lectures on DNA, RNA and biotechnology (pp. 9-
13).
 Inducer. (2015, August 11). Retrieved Saturday, April 16 from
Wikipedia: https://en.wikipedia.org/wiki/Inducer.
 Kovach, T. K. (2014, February 26). Jacob-Monod: The lac operon. In Gene control. Retrieved
from https://www.khanacademy.org/test-prep/mcat/biomolecules/gene-control/v/jacob-monod-
the-lac-operon.
 OpenStax College, Biology. (2015, September 29). Prokaryotic gene regulation. In OpenStax
CNX. Retrieved from http://cnx.org/contents/GFy_h8cu@9.87:drSgdNIj@5/Prokaryotic-Gene-
Regulation.
 Reece, J. B., Urry, L. A., Cain, M. L., Wasserman, S. A., Minorsky, P. V., and Jackson, R. B.
(2011). Bacteria often respond to environmental change by regulating transcription.
In Campbell biology (10th ed., pp. 361-365). San Francisco, CA: Pearson.
 Griffiths AJF, Gelbart WM, Miller JH. 1999. Modern Genetic Analysis. New York: W. H.
Freeman and Company.
Lac Operon Mechanism