Download Discussion of control of the lac operon and mutational analysis

The lac Operon
Laboratory
Purpose: Learning to analyze an example of gene expression
Example: lac operon
Type of control: negative inducible
Today’s Laboratory

Work on mutational analysis problem of lac
operon; page 3-24
Lac Operon:
Negative Control and Induction
Repressor has two binding sites.
Lactose
binding site
DNA binding site
The promoter needs to be in an ideal
conformation for RNA polymerase.
An overview of regulation of the lac operon
Let review a few terms and then look at
the lab problem (table) on page 3-24
important terms of DNA control.
3 important terms of DNA control.



Constitutive activity
Trans acting
Cis acting
Examples of cis and trans acting elements
Cis acting:
c
O mutation
Trans acting
I
mutation
Trans acting:
s
I
mutation
Laboratory Exercise
Haploid and Heterozygous Diploid Mutants


Predict if you expect to obtain:
transcription/translation of the lacZ and lacY genes
when lactose is absent or present for the E.coli.
Try out your ability to reason
through the questions.
PROTEIN
PRODUCT
Β-galactosidase
No Lactose
I+P+O+Z+Y+
I-P+O+Z+Y+
I+P-O+Z+Y+
I+P+OcZ+Y+
I+P+O+Z-Y-/ I-P+O+Z+Y+
I+P+OcZ+Y-/ I+P+O+Z-Y+
I+P+OcZ-Y+/ I-P+O+Z+YISP+O+Z+Y-/ I-P+O+Z-Y+
I+P-OcZ+Y+/ I+P+O+Z-Y-
-
Lactose
+
Permease
No lactose
-
Lactose
+
The next slides are a “key” to the mutations.
Experimental tests on the repressor:

I+ cells synthesize full levels of the lac enzymes only in the
presence of an inducer

I- mutations NO DNA BINDING ACTIVITY, TRANSCRIPTION
OCCURS EVEN IN ABSENCE OF INDUCER!

Is mutation NO LACTOSE BINDING ACTIVITY but has DNA
BINDING ACTIVITY. NO TRANSCRIPTION EVEN WITH
INDUCER!
Experimental mutations to the
repressor
I s=inducer binding region mutated
Inhibits transcription, even with inducer
Lactose
binding site
DNA binding site
I - = DNA binding site mutated,
prevents binding, allows transcription
The End.