Download Chapter 9 homework due 3/31/08 1a. Will lacZ be transcribed and

Chapter 9 homework
due 3/31/08
1a. Will lacZ be transcribed and translated if E. coli are grown in media with only lactose
as a sugar source? YES
What factors will be bound to the lacO and lacP sites these conditions and why? AT P,
the CRP-cAMP COMPLEX WILL BE BOUND + RNA POLYMERASE.
NOTHING WILL BE BOUND TO O.
b. Will lacZ be transcribed and translated if E. coli are grown in media with only glucose
as a sugar source? NO
What factors will be bound to the lacO and lacP sites these conditions? Repressor will
be bound at O.
c. Will lacZ be transcribed and translated if E. coli are grown in media with both lactose
and glucose as sugar sources? NO
What factors will be bound to the lacO and lacP sites these conditions and why? Nothing
will be bound. cAMP levels will be low, so CRP-cAMP complex will not form or
bind to P and neither will RNA Pol. Repressor will be inactivated by binding to
lactose so it will not be bound to O.
d. Will lacZ be transcribed and translated if E. coli are grown in media with only glycerol
as a sugar source? NO
What factors will be bound to the lacO and lacP sites these conditions?
Repressor will be bound to O. CRP-cAMP will be bound to P, but because
repressor is bound to O, RNA Pol won’t have access to the promoter.
2a. Will the trp structural genes be transcribed and translated when E. coli are grown in
media supplemented with high amounts of the amino acid tryptophan? NO
What factors will be bound to the trpO and trpP sites? Tryptophan (the thing the trp
operon enzymes make) will bind to aporepressor to make active repressor, which
will bind to O and prevent RNA Pol from binding to P.
b. Will the trp structural genes be transcribed and translated when E. coli is grown in low
to moderate concentrations of tryptophan?
The levels of trp will not be high enough to turn all aporepressor into active
repressor so some transcripts will begin. There will not be enough active repressor
to ensure that repressor is bound to operator at all times. Therefore transcription
will begin. However, transcription will be attenuated, so the structural genes will
not be translated.
Explain…. Under low to moderate trp concentrations, there will be sufficient
charged trp-tRNAs that the ribosome does need to pause when it gets to the place in
the leader where it should insert two adjacent trps. Thus, the 3-4 stem loop region
Chapter 9 homework
due 3/31/08
will fold into an attenuator structure and the RNA Pol will fall off the DNA… The
mRNA will be attenuated.
3. Will the trp structural genes be transcribed and translated if mutations prevent the 3-4
stem loop in the trpA region from forming? Yes, provided there is not so much trp
around to activate the aporepressor.
4. Why is attenuation a good strategy of gene control for operons that encode the
enzymes that synthesize amino acids?
Because the exact concentration of the end product regulates the level of mRNA
synthesis, the levels of product can be finely regulated by the conditions. Whenever
aa concentrations rise a bit, the relative amount of transcripts that reach full length
drop; but whenever the aa levels drop, full length transcription goes right back up.
5a. What are the similarities and differences between positive and negative control of
transcriptional regulation?
Positive control means that the default mode of a gene is OFF unless it gets the help
of a transcriptional activator. Negative control means that the default mode of a
gene is ON unless a repressor protein comes along and actively prevents RNA Pol
from binding. In both cases something binds to promoter region’s DNA to regulate
transcription—either positively or negatively.
b. What are the similarities and differences between inducible and repressible forms of
negative control?
These are the same in that the gene is on unless a repressor protein binds and turns
the operon off. Also, at least in the cases we studied (lac and trp) the repressor
proteins are made constitutively. They differ in that for the lac operon the
repressor alone is sufficient to bind to DNA and block transcription. The operon is
induced when lactose (the inducer) binds to the repressor and prevents it from
binding to operator, lacO. In contrast, the trp aporepressor is inactive until it binds
to tryptophan (the corepressor); the tryptophan-repressor complex binds DNA (at
trpO) and blocks transcription.
6. Why are operons that make enzymes involved in biosynthetic pathways regulated one
way, while operons that make enzymes involved in degradative pathways regulated
differently?
The genes in biosynthetic pathways need to be on when they need to make product.
But. when the end product levels are sufficient to meet the needs of the cell, there is
no need for more synthesis of the product, so the operons slow down or cease
transcription. They are repressible by the end product.
In contrast, the genes that make enzymes that catabolize sugars, only need to be
active when the sugar is present in the environment. It would waste energy (and
Chapter 9 homework
due 3/31/08
lower survival chances) to make the proteins when they aren’t necessary. Thus,
these genes are in the off mode until the sugar (or some derivative of the sugar) is
present to induce (or derepress) them.
6. E. coli have various mutations in their lac operons (shown in left hand column). Will
the LacZ genes be expressed if the “bugs” are grown in the various conditions listed
across the top of the table? Refer to table 11.2 (p. 453) to see how these different sugars
effect cAMP concentrations.
Lac expression is completely dependent on positive control, so if glucose is around
the lac operon will be inactive.
LacZ gene cannot ever be expressed if the Promoter (P) is mutant because RNA
polymerase won’t be able to bind to DNA. It also won’t be expressed if lacZ gene
itself is mutant.
Is binds to O and will not come off in the presence of lactose.
Oc is a mutation in the binding site for repressor. It will affect only the lacZ genes
downstream of it on same DNA.
Genotype of
lac operon
I+P+O+Z+
I+P-O+Z+
I+P+OcZ+
I+P+O+ZIsP+OcZ+
ISP+O+Z+
Lactose only
yes
no
yes
no
Yes
no
Sugar source
Glucose only
Glucose +
lactose
no
no
no
no
no
no
no
no
no
no
no
no
glycerol
no
no
yes
no
yes
no
7. Are the partial diploids listed below inducible with lactose, constitutively expressed or
uninducible? Assume there is no glucose around…
O+Z+/OcZ+_____constitutive
Is/I+_______ never expressed
OcZ-/O+Z+____inducible____
ISOcZ+/I-O+Z-+____constitutive_____
OcZ+/O+Z+____constitutive____
I-OcZ-/I+O+Z+_______constitutive___
8.Define the terms listed at the end of each chapter covered.