Lac Operon
Lecture 1
Importance of Mutations
• Key tools of genetic analyses
• Allow for fundamental biological processes to be dissected to understand
overall system
Lac Operon Model
• Jacob and Monod
• Deduced structure + function of lac operon via genetic approach
• Analysed interactions of mutations in e.coli that interfered with normal
regulation of lactose metabolism
• Demonstrated operon as basic unit of transcriptional control in bacteria
o Structural genes
o Regulatory genes
o Regulatory DNA sequences
Operon = group of genes physically linked to the chromosome under the control of
the same promotor giving rise to polycistronic mRNA
Regulon = group of genes needed for the same process but located on different
parts of the chromosome with their own promotor, but the promotors are regulated
in the same fashion by the same elements to allow for coordinated expression
Structural Genes
• lacZ codes for B-galactosidase
o major reaction = lactose à glucose and galactose
o minor reaction = lactose à allolactose (inducer) (à inducer binds to
repressor)
• lacY codes for B galactoside permease
, o transports lactose into cell
• LacA codes for B galactoside acetylase
o Unknown function
Regulatory Genes/elements
• Lacl codes for lac repressor (binds to operator) à allosteric protein
o Negative regulator protein
• Operator
• promotor
*lac operon = polycistronic
• One promotor and 1 regulator transcribing 1 mRNA containing several genes
o Translated into many proteins
o Coordinated induction/repression
Negative control of an inducible gene Positive control of an inducible gene
repressor protein binds to operator to Inducer binds to activator protein to allow
prevent transcription in the absence of transcription
lactose
Trans and Cis Acting Control Elements
• Trans-acting control
o Via proteins
o LacL repressor protein
§ Physically linked to lac operon but transcribes independently to
LacZYA and binds to operator
o Catabolite activator protein
§ Located on a different part of the chromosome and transcribed
independently of LacZYA
§ Must bind to cAMP before binding to CAP
• Cis acting control elements
o lacP
§ promotor region
§ binding site of RNA pol
o LacO
§ Operator. Region
§ Binding site of Lac repressor
o CAP site
§ Binding site of CAP + cAMP
Induction of the Lac Operon
• Operon = inducible (default = off in the absence of lactose)
• Lactose = taken up by permease (LacY)
• Lactose is converted to allolactose by B galactosidase
o Allolactose = isomer of lactose
• Interacts with bound lac repressor protein
o Allosteric modification of repressor so that it no longer binds to LacO
o No longer interferes with RNA pol binding to lacP
, o Transcription of operon occurs
• Gene products produces at high levels till lactose depleted
• Inducing signal then drops and the lac repressor is no longer inactivated so it
can bind to lacO again
• Operon is repressed
• Induction = process by which a specific molecule stimulates synthesis of a
given protein
o Molecule responsible for stimulating production of the protein =
inducer
Lac Operon Regulation
• Negative and positive control based on presence and absence of glucose
Negative Control Positive Control à massive increase in transcription
Regulated by lac repressor which binds to Regulated by catabolite activator protein (CAP)
Operator in the presence of GLUCOSE which binds to CAP site via cAMP in the absence
of GLUCOSE
High glucose à adenylate cyclase is inactivated LACTOSE AND NO GLUCOSE
and ATP does not become cAMP
Low glucose à high activity of adenylate cyclase
therefore ATP à cAMP
cAMP + CAP = active form of CAP
active CAP binds to CAP site and interacts with
RNA pol to enhance transcription
*inducer (allolactose) is present, active adenylate
cyclase and high cAMP
Transcription occurs because RNA pol binding to
promotor is enhanced by CAP + repressor is
inactive
LOTS of expression/transcription of the operon
NO LACTOSE AND NO GLUCOSE
Inducer is absent, active adenylate cyclase high
cAMP but repressor is present to block
transcription
GLUCOSE AND NO LACTOSE
Inducer is absent, inactive adenylate cyclase, no
cAMP
à transcription is blocked by lack of CAP and
presence of repressor
LACTOSE AND GLUCOSE = catabolite repression
Inducer is present, inactive adenylate cyclase (because of glucose) and no cAMP à repressor is
inactive and CAP is inactive à basal expression level (transcription occurs but minimally
Lecture 1
Importance of Mutations
• Key tools of genetic analyses
• Allow for fundamental biological processes to be dissected to understand
overall system
Lac Operon Model
• Jacob and Monod
• Deduced structure + function of lac operon via genetic approach
• Analysed interactions of mutations in e.coli that interfered with normal
regulation of lactose metabolism
• Demonstrated operon as basic unit of transcriptional control in bacteria
o Structural genes
o Regulatory genes
o Regulatory DNA sequences
Operon = group of genes physically linked to the chromosome under the control of
the same promotor giving rise to polycistronic mRNA
Regulon = group of genes needed for the same process but located on different
parts of the chromosome with their own promotor, but the promotors are regulated
in the same fashion by the same elements to allow for coordinated expression
Structural Genes
• lacZ codes for B-galactosidase
o major reaction = lactose à glucose and galactose
o minor reaction = lactose à allolactose (inducer) (à inducer binds to
repressor)
• lacY codes for B galactoside permease
, o transports lactose into cell
• LacA codes for B galactoside acetylase
o Unknown function
Regulatory Genes/elements
• Lacl codes for lac repressor (binds to operator) à allosteric protein
o Negative regulator protein
• Operator
• promotor
*lac operon = polycistronic
• One promotor and 1 regulator transcribing 1 mRNA containing several genes
o Translated into many proteins
o Coordinated induction/repression
Negative control of an inducible gene Positive control of an inducible gene
repressor protein binds to operator to Inducer binds to activator protein to allow
prevent transcription in the absence of transcription
lactose
Trans and Cis Acting Control Elements
• Trans-acting control
o Via proteins
o LacL repressor protein
§ Physically linked to lac operon but transcribes independently to
LacZYA and binds to operator
o Catabolite activator protein
§ Located on a different part of the chromosome and transcribed
independently of LacZYA
§ Must bind to cAMP before binding to CAP
• Cis acting control elements
o lacP
§ promotor region
§ binding site of RNA pol
o LacO
§ Operator. Region
§ Binding site of Lac repressor
o CAP site
§ Binding site of CAP + cAMP
Induction of the Lac Operon
• Operon = inducible (default = off in the absence of lactose)
• Lactose = taken up by permease (LacY)
• Lactose is converted to allolactose by B galactosidase
o Allolactose = isomer of lactose
• Interacts with bound lac repressor protein
o Allosteric modification of repressor so that it no longer binds to LacO
o No longer interferes with RNA pol binding to lacP
, o Transcription of operon occurs
• Gene products produces at high levels till lactose depleted
• Inducing signal then drops and the lac repressor is no longer inactivated so it
can bind to lacO again
• Operon is repressed
• Induction = process by which a specific molecule stimulates synthesis of a
given protein
o Molecule responsible for stimulating production of the protein =
inducer
Lac Operon Regulation
• Negative and positive control based on presence and absence of glucose
Negative Control Positive Control à massive increase in transcription
Regulated by lac repressor which binds to Regulated by catabolite activator protein (CAP)
Operator in the presence of GLUCOSE which binds to CAP site via cAMP in the absence
of GLUCOSE
High glucose à adenylate cyclase is inactivated LACTOSE AND NO GLUCOSE
and ATP does not become cAMP
Low glucose à high activity of adenylate cyclase
therefore ATP à cAMP
cAMP + CAP = active form of CAP
active CAP binds to CAP site and interacts with
RNA pol to enhance transcription
*inducer (allolactose) is present, active adenylate
cyclase and high cAMP
Transcription occurs because RNA pol binding to
promotor is enhanced by CAP + repressor is
inactive
LOTS of expression/transcription of the operon
NO LACTOSE AND NO GLUCOSE
Inducer is absent, active adenylate cyclase high
cAMP but repressor is present to block
transcription
GLUCOSE AND NO LACTOSE
Inducer is absent, inactive adenylate cyclase, no
cAMP
à transcription is blocked by lack of CAP and
presence of repressor
LACTOSE AND GLUCOSE = catabolite repression
Inducer is present, inactive adenylate cyclase (because of glucose) and no cAMP à repressor is
inactive and CAP is inactive à basal expression level (transcription occurs but minimally
