COMPREHENSIVE
EXAMINATION
IN
BIOCHEMISTRY
AND MOLECULAR BIOLOGY
PAPER #1
Thursday, January 29, 2004
9:00 a.m. to 11:00 am
PLEASE USE THE
FOLLOWING INSTRUCTIONS:
1.
WRITE YOUR NAME OR I.D. NUMBER ON THE UPPER RIGHT HAND CORNER OF EACH
PAGE.
2.
NUMBER ALL PAGES.
3.
EACH QUESTION SHOULD BE ANSWERED ON A SEPARATE SHEET OF PAPER.
4.
WRITE THE PAPER NUMBER AND QUESTION NUMBER ON EACH PAGE.
5.
USE PEN.
6.
STAPLE PAGES FOR EACH QUESTION SEPARATELY.
TWO-HOUR QUESTIONS:
Q1.
Myxococcus
Q2.
The level of expression of a
protein in a eukaryotic cell can be regulated at many different levels. List the
steps where (from DNA to protein) such regulation is possible and give an
example, describing the mechanism of regulation, in each case.
Q3.
Receptor tyrosine kinases
have been implicated in both normal and neoplastic cell functions.
Describe the mechanisms involved in activating these kinases and
transmitting signals through them from the exterior of a cell to the nucleus to
activate gene expression. Include in
your discussion
a)
a brief
description of ligands for the
receptors
b)
a diagram of
the domain structure of the receptor, labeled with site of ligand binding
and domains involved in the transduction of signal
c)
a discussion of
the initial stages of
activation of the
receptor, comparing homodimeric and
heterodimeric mechanisms
d)
the molecular
basis or the pleiotypic
activation
of the various pathways involved in preparing the cell for
mitogenesis)
e)
a diagram of
the essential protein components
of the central or
canonical mitogenic pathway.
For each component indicate the
class of protein and its basic function(s),
how it is activated, and its
function in the transduction of signal
which results in altered transcription.
Your diagram should
show the ordered recruitment of the
pathway proteins into the signaling complex, and your discussion
should include a description of how each
protein in the pathway is activated.
OR
Q4.
Over the past several years, detailed
crystallographic structures have become available for the small and the large
subunits of the ribosome.
These have greatly expanded our understanding of
RNA structure and RNA-protein interactions.
Describe the general features of RNA structure
and RNA-protein interactions (give specific examples). Are these features also
seen in DNA and DNA-protein structures?
COMPREHENSIVE
EXAMINATION
IN
BIOCHEMISTRY
AND MOLECULAR BIOLOGY
PAPER #2
Thursday, January 29, 2004
1:00 p.m. to 3:00 p.m.
PLEASE USE THE
FOLLOWING INSTRUCTIONS:
1.
WRITE
YOUR NAME OR I.D. NUMBER ON THE UPPER RIGHT HAND CORNER OF EACH PAGE.
2.
NUMBER ALL PAGES.
3.
EACH QUESTION SHOULD BE
ANSWERED ON A SEPARATE SHEET OF PAPER.
4.
WRITE THE PAPER NUMBER
AND QUESTION NUMBER ON EACH PAGE.
5.
USE PEN.
6.
STAPLE PAGES FOR EACH
QUESTION SEPARATELY.
ANSWER ONE OF
THE FOLLOWING TWO QUESTIONS (30 MINUTES)
Q5.
A temperature-sensitive mutant of
E.coli was found to immediately shut off protein synthesis upon
elevating the temperature to 42 0C. Analysis of the cells after the
shutoff revealed that aminoacyl-tRNAs were fully charged, that the P-site of
ribosomes contained peptidyl-tRNAs of many lengths, and that the ribosome A-site
was empty.
(i) what is the likely defect
in the mutant strain? Provide a brief description of how you arrived this
conclusion.
(ii) devise an experiment that could be used to test whether your
conclusion is correct.
Q6.
Many
eukaryotic mRNAs contain untranslated regions at their 5’ and 3’ ends (UTRs).
What is known about the function of these UTRs? Give examples.
ANSWER ONE OF
THE FOLLOWING TWO QUESTIONS (30 MINUTES)
Q7.
The tumor suppressor gene
p53 is found mutated in a number of different tumors.
Describe its role in the cellular response to DNA damage, and discuss how
mutations in this gene could lead to tumor progression.
OR
Q8.
What is a dominant negative mutation?
How do you explain the phenotype of such mutations?
Dominant negative mutations are found in the gene for the cellular
regulatory protein p53 in DNA from many tumors.
Explain why this type of mutation is selected.
ANSWER ONE OF
THE FOLLOWING TWO QUESTIONS (30 MINUTES)
Q9.
Chromatin structure.
a.
Discuss the
structure of the core nucleosome in terms of the following: the structure of the
histones and how the histones associate with each other, DNA organization,
DNA-histone contacts, types of DNA preferred for nucleosome assembly.
b.
You’ve just
reconstituted chromatin in vitro. What method do you use to determine whether
nucleosomes were formed and whether the nucleosomes were spaced appropriately?
Describe what the results should look like for properly reconstituted chromatin.
What are the two main types of chromatin altering
activities involved
Q10.
The only function of folate coenzymes in the body appears to be mediating the transfer of one-carbon units. Folate coenzymes act as acceptors and donors of one-carbon units in a variety of reactions critical to the metabolism of nucleic acids and amino acids.
Nucleic acid metabolism:
Folate coenzymes play a vital role in DNA metabolism through two different
pathways.1) The synthesis of DNA from its precursors is dependent on folate
coenzymes. 2) A folate coenzyme is required for the synthesis of methionine, and
methionine is required for the synthesis of S-adenosylmethionine (SAM). SAM is a
methyl group (one-carbon unit) donor used in many biological methylation
reactions, including the methylation of a number of sites within DNA and RNA.
Methylation of DNA may be important in cancer prevention.
Do you agree with the
statements in the last paragraph? If so, state why. If you do not agree with the
statements, explain why and rewrite the paragraph into one that you consider
correct.
Whether you agree or not,
make a list of compounds that require methylation. In the case of DNA and RNA be
specific about what is methylated.
Q11.
Compare the mechanism of DNA
replication initiation in prokaryotes and eukaryotes highlighting the
differences and similarities where they exist.
Include in your discussion the following topics, (1) origin structure and
function, (2) initiator proteins, (3) the role of DNA polymerases I and III (in
bacteria), and Pol a,
d, and
e (in eukaryotes).
OR
Q12.
RNA polymerases carry out
transcription of DNA into RNA in all cells.
a)
In prokaryotes,
how does the RNA polymerase find the start site for a gene and the direction for
transcription.
b)
How does the
polymerase know when to stop.
c)
Are these
mechanisms the same for eukaryotes, for Archaea ?
(END OF PAPER #2)
COMPREHENSIVE
EXAMINATION
IN
BIOCHEMISTRY
AND MOLECULAR BIOLOGY
PAPER #3
9:00 a.m. to 11:00 a.m.
PLEASE USE THE
FOLLOWING INSTRUCTIONS:
1.
WRITE
YOUR NAME OR I.D. NUMBER ON THE UPPER RIGHT HAND CORNER OF EACH PAGE.
2.
NUMBER ALL PAGES
3.
EACH QUESTION SHOULD BE
ANSWERED ON A SEPARATE SHEET OF PAPER
4.
WRITE THE PAPER NUMBER
AND QUESTION NUMBER ON EACH PAGE.
5.
USE PEN.
6.
STAPLE PAGES FOR EACH
QUESTION SEPARATELY.
ANSWER ONE OF
THE FOLLOWING TWO QUESTIONS (30 MINUTES)
Q13.
A wide range of approaches
has been developed to characterize conformational or dynamical states of native
proteins in atomic detail by NMR spectroscopy, and evidence for the contribution
of conformational entropy towards the stability of protein-ligand complexes has
been reported.
a)
List and describe
two different NMR-relaxation parameters that are used to characterize the
dynamical properties of backbone amide groups of residues in a protein.
b)
Describe how
changes in a protein's dynamical properties or "conformational entropy" could
contribute to the stability of a given protein-ligand complex.
Q14.
A wide range of approaches
has been developed to characterize nonnative states of proteins in atomic detail
by NMR spectroscopy, and evidence for the persistence of native-like structure
even under strongly denaturing conditions has been reported.
a)
List and describe
two different NMR-measurable parameters that indicate that the backbone amide
group of a given residue or stretch of residues in a protein behaves either as a
random coil or is restricted to a given residual structure.
b)
Describe how
enthalpy and entropy could contribute to the observed persistence of long-range
structure under denaturing conditions.
ANSWER ONE OF
THE FOLLOWING TWO QUESTIONS (30 MINUTES)
Q15.
Taking into consideration the large number of hydrogen bonds, salt bridges and favourable Van der Waal’s interactions between atoms in a protein molecule, why are most proteins so unstable ? In other words, explain why the difference in free energy between the folded and unfolded states of a protein molecule in aqueous solution is typically small (3 to 5 Kcal per Mole).
Q16.
Describe in detail the catalytic mechanism of one named enzyme. Indicate the substrate(s) for the enzyme, the product(s) and the nature of the transition state intermediate. Indicate the roles of any specific amino acids that participate in catalysis and describe how the transition state of the reaction is stabilized by the enzyme
ANSWER ONE OF
THE FOLLOWING TWO QUESTIONS (30 MINUTES)
Q17.
Briefly describe the structural and functional significance of
each of the following protein domains or motifs.
a.
SH2 domain
b.
Zn++ finger
c.
CAAX motif
d.
SH3 domain
Q18.
ANSWER ONE OF
THE FOLLOWING TWO QUESTIONS (30 MINUTES)
Q19.
Draw labeled diagrams of the Lineweaver Burke plots for (a) competitive and (b) non-competitive inhibitors. Explain why only one of these two types of inhibitors affects the KM for substrate, while the other does not. For one named enzyme give an example of a competitive inhibitor and explain how it is chemically similar to the substrate.
Q20.
a) (5 min) Consider the
replication of a 2 million bp chromosome containing a
single, bidirectional
origin (similar to that of
E. coli). If it takes
1 sec for any polymerase to bind
at a primer-template complex,
replisomes synthesize DNA at 1,000 bp per sec (the rate of replication fork
travel with leading + lagging strand synthesis), and the average
processivity of the replisome polymerases is 100,000 polymerization
reactions (turnovers) per primer-template binding event,
how long will it take to duplicate the chromosome? You may assume
that the diffusion rate and synthesis rates are the only rate-limiting steps in
this process. SHOW YOUR WORK for full
credit!
b) (5 min) What is the
dwell time of
each replisome polymerase?
c) (5 min) Ok, now
consider the same scenario, except substitute the replisome polymerase with a
repair polymerase that has an average
processivity of 1000 bp and a
synthesis rate of 10 bp per sec. How
long will it take the replisomes with the repair polymerases to duplicate the
chromosome?
d) (5 min) What is the
dwell time of
each repair polymerase?
e) (10 min) Briefly
comment on the relative dwell times of the
replisome polymerase and repair polymerase that you calculated, and on the role
of turnover rates and dwell times to the processivity of each polymerase in
these scenarios. Based on your calculations, do you think the repair polymerase
in this example employed the sliding clamp of the replisome or not?
STATE YOUR LOGIC for full credit.
(END OF EXAM)