the lactose (lac) operon - an example for prokaryotic gene regulation

The lactose (lac) operonThe lactose (lac) operon

- an example for prokaryotic gene regulation- an example for prokaryotic gene regulation

19.

LCT and MCM 6 genesLCT and MCM 6 genes

Breakdown of lactosein human

Breakdown of lactosein human

Nobel prize for the discovery of lac operonNobel prize for the discovery of lac operon - 1965- 1965

Francois Jacob Jacques MonodFrancois Jacob Jacques Monod

Pasteur Institute, Paris, FrancePasteur Institute, Paris, France

1.

DNA lacZ

repressor CAP site RNA pol operator structural genes terminator

lacI O lacY

lacA

C E

promoter

P

Cytoplasm cell wall outer world

promoter Structural genes DNAT

catabolit activator protein binding site

E. coli DNA

binding site

operon

The structure of lac operonThe structure of lac operon2.

Structure lac operonStructure lac operon 3.

DNA DNAlacZ lacI O lacY

lacA

C E P

repressor CAP site entry site operator structural genes terminator

T

polrep

RRepressionepressionRRepressionepression

rep: repressorpol: RNA polymerase

4.

DNA DNAlacZ lacI O lacY

lacA

C E P

rep

lactose

galactose

glucose

repressor CAP site entry site operator structural genes terminator

T

rep: repressorpol: RNA polymerase

5.

InductionInductionInductionInduction

DNA DNAlacZ lacI O lacY

lacA

C E P

trans-acetylase

permease

rep

b-gal

repressor CAP site entry site operator structural genes terminator

T

pol

rep: repressorpol: RNA polymerase

5.

InductionInductionInductionInduction

DNA DNAlacZ lacI O lacY

lacA

C E P

CA

P

cAMP No glucose

repressor CAP site entry site operator structural genes terminator

T

CAP: catabolite activator protein

6.

ActivationActivationActivationActivation

DNA DNAlacZ lacI O lacY

lacA

C E P

cAMPNo glucose

repressor CAP site entry site operator structural genes terminator

T

CA

P

pol

6.

ActivationActivationActivationActivation

DNA DNAlacZ lacI O lacY

lacA

C E P

CA

P

pol

b-galpermease

trans-acetylase

repressor CAP site entry site operator structural genes terminator

T

6.

ActivationActivationActivationActivation

DNA DNAlacZ lacI O lacY

lacA

C E P

No cAMPGlucose is present

repressor CAP site entry site operator structural genes terminator

T

6.

CA

P

pol

catabolite repression (endproduct inhibition)

ActivationActivationActivationActivation

DNA DNAlacZ lacI O lacY

lacA

C E P

CA

P Combined regulationCombined regulationpol

cAMP

glucose

lactose

rep

b-galpermease

ATP

adenylate cyclase lactose

allolactose (inductor!)

Glucose -Glucose -

Lactose +Lactose +trans-acetylase

Structure and operation of lac operonStructure and operation of lac operon

repressor CAP site entry site operator structural genes terminator

T

7.

adenylate cyclase

cyclic AMP

DNA DNA lacI O lacY

lacA

C E P

CA

P

cAMPNo glucose

lacZ

polrep

Glucose -Glucose -

Lactose +Lactose +

repressor CAP site entry site operator structural genes terminator

T

Mutant beta-galMutant beta-gal8.

DNA DNA lacI O lacY

lacA

C E P

CA

P

permease

lacZ

pol

lactose

b-gal*

Glucose -Glucose -

Lactose +Lactose +trans-acetylase

repressor CAP site entry site operator structural genes terminator

T

8. Mutant beta-galMutant beta-gal

DNA DNAlacZ lacI O lacY

lacA

C E P

CA

P

pol

rep

b-galpermease

Glucose -Glucose -

Lactose +Lactose +

repressor CAP site entry site operator structural genes terminator

T

transcription stop

stop

Transcription stop in genesTranscription stop in genes9.

DNA DNAlacZ lacI O lacY

lacA

C E P

CA

P

pol

rep

b-gal

Glucose -Glucose -

Lactose +Lactose +

repressor CAP site entry site operator structural genes terminator

T stop

Transcription stop in genesTranscription stop in genes9.

DNA DNAlacZ lacI O lacY

lacA

C E P

CA

P

pol

rep

Glucose -Glucose -

Lactose +Lactose +

repressor CAP site entry site operator structural genes terminator

T stop

Transcription stop in genesTranscription stop in genes9.

DNA DNAlacZ lacI Oc lacY

lacA

C E P

permeaseb-gal

Glucose -Glucose -

Lactose -Lactose -trans-acetylase

The operator is not able to bind the repressor

repressor CAP site entry site operator structural genes terminator

T

CA

P

polrep

pol

The Oc mutation of operonThe Oc mutation of operon10.

DNA DNAlacZ O lacY

lacA

C E P

permeaseb-gal

Glucose -Glucose -

Lactose -Lactose -trans-acetylase

The repressor is not able to bind to the operator

repressor CAP site entry site operator structural genes terminator

T lacI

CA

P

polrep

pol

The I- mutation of repressorThe I- mutation of repressor11.

DNA DNAlacZ O lacY

lacA

C E P

lacI*

Glucose -Glucose -

Lactose +Lactose +

The repressor is unable to bind lactose

repressor CAP site entry site operator structural genes terminator

T

CA

P

pol

pol

rep

The Is mutation of repressorThe Is mutation of repressor12.

DNA DNAlacZ lacI O lacY

lacA

C E P

CA

P

cAMP No glucose

CAP is unable to bind cAMP

Glucose -Glucose -

Lactose +Lactose +

repressor CAP site entry site operator structural genes terminator

T

Mutant CAP-IMutant CAP-I13.

DNA DNAlacZ lacI O lacY

lacA

C E P

cAMP

Glükóz -Glükóz -

Laktóz +Laktóz +

No glucose

repressor CAP site entry site operator structural genes terminator

T

pol

CA

P

Mutant CAP-IMutant CAP-I13.

CAP is unable to bind cAMP

DNA DNAlacZ lacI O lacY

lacA

C E P

CAP can bind promoter in the absence of cAMP

Glucose +Glucose +

Lactose +Lactose +

b-gal

trans-acetylase

rep

permease

repressor CAP site entry site operator structural genes terminator

T

CA

P

pol

pol

Mutant CAP-IIMutant CAP-II14.

Too many repressorsToo many repressors

DNA lacZ lacI O lacA

C E P

Repressor Repressor – on the plasmid– on the plasmid

repressor CAP site entry site operator structural genes terminator

DNA lacI P

Glucose -Glucose -

Lactose +Lactose +

DNAT

CA

P

polrep

rep

rep

rep

lacY

plasmid

DNA lacI P

plasmid

DNA lacI P

plasmid

DNA lacI P

plasmid

rep

rep

reprep

15.

DNA lacZ lacI O lacA

C E P

Operator Operator – on the plasmid– on the plasmid

represssor CAP site entry site operator structural genes terminator

DNA

Glucose -Glucose -

Lactose -Lactose -

DNAT

CA

P

lacY

plasmid O

DNA plasmid O

DNA plasmid O

DNA plasmid O

rep

rep

rep

b-galtrans-acetylasepermease

pol

pol

Too many Too many operatorsoperators16.

DNA DNSlacZ lacI Oc lacY-

lacA

C E P

b-gal

Glucose -Glucose -

Lactose -Lactose -

OOcc mutation mutation – on bacterial DNA– on bacterial DNA

perm*

T

rep

CA

P

pol

pol

trans-acetylase

17a. Partial diploidsPartial diploids: cis or trans-effect?: cis or trans-effect?Partial diploidsPartial diploids: cis or trans-effect?: cis or trans-effect?

DNA F plasmidlacZ- lacI O lacY

lacA

C E P

DNA DNAlacZ lacI Oc lacY-

lacA

C E P

CA

P

CA

P

polb-gal

Glucose -Glucose -

Lactose -Lactose -trans-acetylase

OOcc mutation mutation – on bacterial DNA– on bacterial DNA

permeaseb-gal*trans-acetylase

perm*

Glucose -Glucose -

Lactose +Lactose +

repressor CAP site entry site operator structural genes terminator

T

T

rep

reppol

pol

polrep

Coli lacOc lacZ+ lacY- -gal: continuously expressedF’ lacO+ lacZ- lacY+ permease: only with lactose

mutation

17b. Partial diploidsPartial diploids: cis or trans-effect?: cis or trans-effect?Partial diploidsPartial diploids: cis or trans-effect?: cis or trans-effect?

DNA lacZ lacI* O lacY-

lacA

C E P

II-- mutation mutation – on bacterial DNA– on bacterial DNA

DNA F plasmid lacZ- lacI O lacY

lacA

C E P

repGlucose -Glucose -

Lactose -Lactose -

Glucose -Glucose -

Lactose +Lactose +

b-gal

trans-acetylase

perm*

permeaseb-gal*

repressor CAP site entry site operator structural genes terminator

DNAT

T

CA

P

CA

P

polpol

rep

rep

rep

rep

rep

trans-acetylase

Coli lacI- lacO+ lacZ+ lacY- -gal with lactose

F’ lac I+ lacO+ lacZ- lacY+ permeáz with lactose

mutation

18. Partial diploidsPartial diploids: cis or trans-effect?: cis or trans-effect?Partial diploidsPartial diploids: cis or trans-effect?: cis or trans-effect?