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Name _______________________________________________
Biochemistry Biol 100 Summer 2014
Jeremy Lee Eryn Wicklow
Midterm Exam July 10, 2014
BE SURE TO PUT YOUR NAME ON ALL PAGES. (14 PAGES TOTAL) The
following exam is composed of four parts (112 points total:) Part
1. Multiple-choice (28 points total); Part 2. Fill-in (24 points
total); Part 3. Calculation Problem (12 points); Part 4. Written
Answer (48 points total.) Part 1. Multiple-choice. Choose the one
best answer and fill-in the corresponding bubble
on the Scantron form. (1 point each) 1. Proteins that are to be
utilized in the plasma membrane are most commonly A. synthesized on
free ribosomes in the cytosol. B. synthesized on ribosomes in the
rough ER. C. transported to the plasma membrane in secretory
vesicles that bud off from the rough ER. D. A and C E. B and C
ANSWER: B 2. Hydrophobic molecules tend to aggregate in aqueous
solutions because A. the hydrophobic molecules are negatively
charged so are repelled by the positively charged water molecules.
B. the hydrophobic molecules are positively charged, so are
repelled by the negatively charged water molecules. C. this
decreases the entropy of water molecules surrounding the
hydrophobic molecules. D. the hydrophobic molecules bind only to
the positive charges of polar water molecules. E. this increases
the entropy of water molecules surrounding the hydrophobic
molecules. ANSWER: E 3. Acetic acid has a pKa of 4.76, whereas
pyruvic acid has a pKa of 2.50. Which of the following is/are true?
A. Pyruvic acid is a stronger acid than acetic acid. B. The pH at
which acetic acid is fully dissociated is 4.76. C. The pH at which
acetic acid is half dissociated is 4.76. D. A and B E. A and C
ANSWER: E
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Name _______________________________________________ 2 4. A
characteristic of a buffer is that it A. is a strong acid. B. is a
weak acid. C. prevents large changes in pH, even with a significant
input of protons. D. A and C E. B and C
ANSWER: E 5. At neutral pH (pH=7), an amino acid typically has
A. just its amino group protonated. B. just its carboxyl group
protonated. C. both its amino group and its carboxyl group
deprotonated. D. both its amino group and its carboxyl group
protonated. ANSWER: A 6. An important characteristic of the amino
acid cysteine is A. its side chain has a sulfhydryl group. B. it
can form a covalent bond with another cysteine. C. its side chain
is hydrophobic. D. A and B E. all of the above ANSWER: D 7. At
physiological pH, histidine can be A. negatively charged. B.
positively charged. C. uncharged. D. A and C E. B and C ANSWER: E
8. In a polypeptide, rotation does not occur around which bond? A.
-carbon to amino nitrogen bond. B. -carbon to carbonyl carbon bond
C. peptide bond D. A and B E. all of the above ANSWER: C 9. In an
alpha helix, the amino acid side chains are oriented A. toward the
inside of the helix. B. toward the outside of the helix. C.
parallel to the long axis of the helix. D. randomly with respect to
the helix. E. all of the above ANSWER: B
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Name _______________________________________________ 3 10. In
proteins, a difference between a beta sheet composed of two
antiparallel -strands and a beta sheet composed of two parallel
strands is that in a beta sheet composed of antiparallel strands A.
there are fewer hydrogen bonds between amino acids on the two
strands. B. each amino acid on a strand forms one hydrogen bond
with each of two amino acids on the other strand. C. each amino
acid on a strand forms two hydrogen bonds with only one amino acid
on the other strand. D. A and B E. A and C
ANSWER: C 11. Major and minor grooves are found in a DNA double
helix because of A. repellency between phosphate groups on the two
strands of DNA. B. the angles of the bonds between the deoxyribose
sugars and the bases. C. the angles between phosphodiester bonds on
adjacent nucleotides on a chain. D. hydrogen bonding between
nucleotides on the same strand of DNA. E. the difference in the
number of hydrogen bonds between G-C pairs and A-T pairs. ANSWER: B
12. In DNA replication, the DNA polymerase catalyzes the reaction
between the 3 OH of one nucleotide and A. the -phosphate of a
deoxyribonucleotide triphosphate. B. the -phosphate of a
deoxyribonucleotide triphosphate. C. the -phosphate of a
deoxyribonucleotide diphosphate. D. the -phosphate of a
deoxyribonucleotide diphosphate. E. the -phosphate of a
deoxyribonucleotide phosphate. ANSWER: B 13. The function of DNA
helicase in DNA replication is to A. unwind the DNA. B. rewind the
DNA. C. maintain DNA polymerases attachment to the template strand.
D. A and B E. all of the above. ANSWER: A 14. The function of the
sliding clamp in DNA replication is to A. prevent single-stranded
DNA from base-pairing to itself. B. unwind DNA. C. increase and
maintain DNA polymerases processivity. D. A and B E. all of the
above. ANSWER: C
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Name _______________________________________________ 4 15. DNA
ligase functions in A. base excision repair B. nucleotide excision
repair C. lagging strand synthesis in DNA replication D. A and B E.
all of the above
ANSWER: E 16. Ultraviolet light damages DNA by creating A.
alkylated bases. B. reduced bases. C. dimers of pyrimidines. D.
oxidized bases. E. deaminated bases. ANSWER: C 17. In E. coli, the
sigma subunit of RNA polymerase is critical for A. binding of the
RNA polymerase to the transcription initiation site. B. unwinding
of the DNA to allow reading of the DNA template. C. binding of the
RNA polymerase to the Shine-Dalgarno sequence of the promoter. D.
binding of the RNA polymerase to the operator sequence. E. binding
of the RNA polymerase to the -10 and -35 sequences. ANSWER: E 18. A
characteristic of RNA polymerase that is not a characteristic of
DNA polymerase is that RNA polymerase A. is not a processive
enzyme. B. does not need a primer to initiate formation of a new
strand. C. reads along the template strand from 3' to 5'. D. A and
B E. all of the above ANSWER: B 19. Post-transcriptional
modification of a newly transcribed prokaryotic tRNA includes A.
addition of a 5 cap. B. addition of a poly A tail. C. addition of
the nucleotides C-C-A to the 3 end D. A and B E. all of the above
ANSWER: C
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Name _______________________________________________ 5 20. When
glucose levels are high in E. coli A. there is more cAMP bound to
CAP protein, decreasing the CAP protein's binding to the lac
operon. B. there is more cAMP bound to CAP protein, increasing the
CAP protein's binding to the lac operon. C. there is less cAMP
bound to CAP protein, decreasing the CAP protein's binding to the
lac operon. D. there is less cAMP bound to CAP protein, increasing
the CAP protein's binding to the lac operon.
ANSWER: C 21. A major function of the TFIIH basal transcription
factor in eukaryotes is to A. bind to the GC box of the promoter.
B. interact with the mediator complex to allow RNA polymerase to
bind to the promoter. C. phosphorylate the C-terminal domain of the
RNA polymerase, allowing it to escape the promoter. D.
phosphorylate the C-terminal domain of TFIID, causing it to release
RNA polymerase from the promoter. E. phosphorylate the C-terminal
domain of histone proteins, making the promoter more available for
binding by RNA polymerase. ANSWER: C 22. In eukaryotic
transcription, enhancers A. bind proteins that interact with the
mediator complex to affect the levels of transcription. B. bind to
basal transcription factors to allow formation of the
pre-initiation complex. C. bind to the spliceosome to facilitate
intron removal. D. bind to the mediator complex to affect levels of
transcription. E. modify histone tails to affect the levels of
transcription. ANSWER: A 23. In spliceosome-mediated removal of an
intron, the second transesterification reaction involves A.
cleavage of the phosphodiester linkage between the nucleotide at
the 3 end of the upstream exon and the nucleotide at the 5 end of
the intron. B. formation of a bond between the guanosine at the 5
end of the intron and the branch site adenosine. C. formation of a
phosphodiester bond between the 3' OH of the nucleotide at the 3
end of the upstream exon and the 5' phosphate of the nucleotide at
the 5 end of the downstream exon. D. A and B E. all of the above
ANSWER: C
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Name _______________________________________________ 6 24. The
first step in "charging" a tRNA with an amino acid (forming a bond
between the tRNA and an amino acid) involves a reaction between the
amino acid and A. the 3' end of the tRNA. B. EF-Tu. C. ATP. D. AMP.
E. the small unit of the ribosome.
ANSWER: C 25. A characteristic of prokaryotic translation that
is not a characteristic of eukaryotic translation is that in
prokaryotic translation A. the mRNA is associated with the small
unit of the ribosome prior to the large subunit of the ribosome
joining the complex. B. the initiator-aminoacyl-tRNA initially
associates with the mRNA at the start codon. C. the complete
ribosome is composed of two subunits. D. the initiator methionine
is not formylated. E. peptide bond formation is a catalyzed by an
rRNA in the large subunit of the ribosome. ANSWER: B 26. In a
reaction at equilibrium, A. G0 D. Can't tell without knowing
concentrations of reactants and products. E. G may be >0 or
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Name _______________________________________________ 7
Part 2. Fill-in. In each blank, write in the appropriate, and
best, word or phrase. (1 point per blank, except where noted.) 1. A
bond that forms between atoms, which involves sharing of an
electron between the two atoms, is called a(n)
___covalent____________________ bond. 2. In eukaryotic cells,
secretory vesicles are produced from an organelle called the
___Golgi complex (or apparatus.) 3. Draw the basic structure of an
amino acid (2 points)
(Note: this is worth 2 points. Also, they don't need to put text
labels.)
___________________________________________________________ 4. Many
membrane proteins, like porins, are said to show inside-out
conformation because ___hydrophobic_______________________ amino
acids tend to be found on the exterior of the functional protein.
5. The bond in a nucleotide that links the base to the sugar is
called a(n) ___N-glycosidic (or -glycosidic) bond (or linkage.) 6.
The two major kinds of interactions that occur between bases in
double-stranded DNA are ___hydrogen_______________________________
bonds and ____Van der Waals interactions (or forces)_. 7. In
eukaryotic DNA, the basic structure of DNA packaging is called a(n)
___nucleosome________________, which is composed of DNA wrapped
around a core of eight ___histone___________________ proteins.
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Name _______________________________________________ 8 8. If an
incorrect nucleotide is added to a growing strand of DNA by DNA
polymerase, the incorrect nucleotide is usually detected by the
____proofreading__________________ activity of the DNA polymerase.
9. In base excision repair, the first step is removal of the
damaged base by an enzyme called a(n) ____DNA
glycosylase______________________________. 10. The following is a
portion of the transcribed region of a gene:
5 ACCTGTCGATGCCCAGTCGTGT 3 (strand #1) 3 TGGACAGCTACGGGTCAGCACA
5 (strand #2)
Here is a corresponding portion of the mRNA that is produced by
transcription of this gene: 5 ACCUGUCGAUGCCCAGUCGUGU 3 With respect
to transcription, DNA strand #1 is the _sense (or
coding)___________ strand and DNA strand #2 is the _anti-sense (or
template)_ strand. 11. In eukaryotes, phosphorylation of the
_C-terminal domain of RNA polymerase_______________ is required for
transcription to be initiated. This phosphorylation is catalyzed by
the basal transcription factor __TFIIH_________________. 12. In
eukaryotes, after cleavage of the pre-mRNA, a(n) _poly(A)
tail_______________ is added to the 3 end by the enzyme ___poly (A)
polymerase___________________. 13. In a spliceosome, the catalytic
activity for intron removal and splicing is a function of the
___snRNA_______________________ molecules within the snRNPs. 14. A
nucleotide that is frequently found at the 5' end of tRNA
anticodons and which can base pair with three different bases on
mRNA is __inosine______________________________. 15. The group of
enzymes that catalyze the reactions that form bonds between tRNAs
and amino acids is called
____amino-acyl-tRNA-synthetases__________________________. 16. The
factor that "escorts" an aminoacyl-tRNA to the ribosome is called
____EF-Tu________________________________. 17. Hydrolysis of
_____GTP____________ provides the energy by which the translation
factor called ___EF-G (or elongation factor-G)____ translocates the
mRNA and the tRNAs along the
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Name _______________________________________________ 9 ribosome
so that the next codon moves into the position to associate with a
new aminoacyl-tRNA.
Part 3. Calculation problem. Complete parts A, B and C. You must
show your work to get credit. Equations you might need are listed
at the bottom of the page. For the following reaction: A B K'eq =
0.2 For this reaction: A. Calculate Go' for the reaction (3
points).
K'eq = e-Go'/RT 0.2 = e-Go'/RT ln 0.2 = ln e-Go'/RT -1.61 =
-Go'/RT -1.61 x 2.47 = -Go' Go' =
__3.98____________________________________ B. Determine G for the
reaction when the initial concentrations of the two compounds are:
[A] = 6 x 10-2 M [B] = 3 x 10-3 M G = Go' + RT ln ([C][D] / [A][B])
G = Go' + RT ln ([products] / [reactants]) G = 3.98 + 2.47 ln
(0.003 / 0.06) G = 3.98 + 2.47 ln (0.05) G = 3.98 + 2.47 x (-3.0) G
= 3.98 7.41 G = -3.43
G = _-3.43______________________________________ (5 points) C.
At the concentrations given in part B, and under standard
conditions (25oC, pH=7,) will the reaction occur spontaneously?
Explain. (4 points) Yes, the reaction will occur spontaneously
since, at those concentrations, G
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Name _______________________________________________ 10 Go = -
RT ln [C][D] / [A][B] (at equilibrium)
Part 4. Written Answer. The following section contains 5
questions. CHOOSE 3 of the 5 and write thorough answers. Only the
first 3 questions you answer will be graded (unless you completely
cross out your answer.) Use the bottom and back of the last page if
you need more room. (16 points each) 1. Describe all the important
functions of each of the following in eukaryotic transcription: RNA
polymerase II; TFIID; TFIIH; activator proteins; enhancers; histone
acetyltransferases; histone deacetylases; chromatin remodeling
engines. In addition, describe how the process of transcription in
terminated in eukaryotes.
RNA polymerase II: Binds promoter sequence; unwinds and rewinds
DNA; reads anti-sense (template) strand of DNA and adds
complementary RNA nucleotides 5' to 3', creating ester bonds
between them, to synthesize pre-mRNA.
TFIID: Basal transcription factor that initiates formation of
the pre-initiation complex by
binding to the promoter (e.g. TBP subunit of TFIID binds to the
TATA box sequence of the promoter.)
TFIIH: Basal transcription factor that induces RNA polymerase to
"melt" (initial unwinding of)
DNA at the promoter. Also, catalyzes phosphorylation of
C-terminal domain (CTD) of RNA polymerase II, allowing it to escape
the promoter and initiate transcription.
Activator proteins: bind enhancer sequences and increase level
of transcription by 1)
interacting with mediator complex to facilitate formation of the
RNA pol II pre-initiation complex; and/or 2) recruitment of
chromatin remodelers and/or histone modifiers that facilitate
binding of basal transcription factors/RNA pol II to promoter.
Enhancers: DNA sequences frequently upstream of promoter (also
may be in introns) to which
activator proteins (or repressor proteins) bind to influence the
level of transcription. Histone acetyltransferases: add acetyl
groups to amino acids (especially lysines) in the N-
terminal tails of core histone proteins of nucleosomes. This
generally "loosens" the association between the core histones and
DNA, facilitating binding of basal transcription factors/ RNA pol
II and activity of chromatin remodelers. Activity generally
associated with increased transcription.
Histone deacetylases. remove acetyl groups from amino acids in
the N-terminal tails of core
histone proteins of nucleosomes. This generally "tightens" the
association between the core histones and DNA, inhibiting binding
of basal transcription factors/ RNA pol II and the activity of
chromatin remodelers. Activity generally associated with decreased
transcription.
Chromatin remodeling engines: modify the interactions between
DNA and associated proteins
(e.g. histones.) For example may "spin off" a segment of DNA
from histone core of nucleosome to make DNA more available for
binding of basal transcription factors and
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Name _______________________________________________ 11 RNA pol
II. (May be associated with increased or decreased transcription,
depending on the type of remodeling.) 2. Address each of the
following with respect to a chemical reaction: A. Explain what
activation energy is with respect to a chemical reaction. B.
Indicate whether the "activation energy" of a chemical reaction is
included in the calculation of G for a reaction; explain. C.
Describe how enzymes catalyze reactions with respect to activation
energy, and draw and explain a graph depicting this effect. D.
Describe the effect that an enzyme has on the G of a reaction.
A. Activation energy is the difference in free energy between
reactant(s) (substrate(s)) and the transition state, the
conformation between the reactant(s) and product(s). In other
words, activation energy is the amount of energy that must be added
to allow the reactant(s) to reach the transition state.
B. Activation energy is NOT included in calculation of G because
all the energy added to
reach transition state is released/recovered when transition
state is transformed into the product(s.) (G is simply the
difference in free energy of products and reactants.)
C. Enzymes catalyze reactions by lowering the activation energy
required to reach the
transition state, thus facilitating formation of the transition
state and increasing the rate of the reaction. As shown in the
graph below, the effect of an enzyme is to lower the free energy of
transition state, meaning that the input of energy to reach
transition state, activation energy, is reduced by the enzyme
Graph should look something like this:
D. Enzymes do NOT affect the levels of free energy in the
reactants or in the products, and since G is the difference in free
energy of products and reactants, enzymes have no effect on G.
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Name _______________________________________________ 12 3.
Describe, in detail, the processing of a eukaryotic pre-mRNA to
produce an mRNA that is ready for translation. Be sure to include
then name and functions of the important molecules involved in the
various types of processing. Also, indicate how this processing
affects the number of proteins that can be encoded by a single
gene. 1. Addition of 5' cap (7-methyl guanosine) to 5' end of
pre-mRNA:
A. The enzyme RNA triphosphatase cleaves off one phosphate group
(gamma phosphate) from the 5' end of the 5' (first) nucleotide of
the pre-mRNA
B. The enzyme guanylyltransferase, creates an ester bond between
the beta phosphate (now the terminal phosphate) of the 5'
nucleotide of the pre-mRNA and the alpha phosphate of an additional
guanosine nucleotide.
C. The enzyme methyltransferase adds a methyl (CH3) group to the
N-7 of the guanosine.
2. Cleavage of pre-mRNA and addition of poly (A) tail.
A. Several (about 20) nucleotides downstream (3') of the signal
sequence (polyadenylation signal) an endonuclease cleaves the
pre-mRNA.
B. poly (A) polymerase adds a large number (tens to hundreds) of
adenosines to the 3' cut end of the pre-mRNA.
3. Removal of introns and splicing of exons.
A. Spliceosome subunits associate with each intron. B. First
transesterification reaction, catalyzed by U2 and U6 snRNAs: Oxygen
of branch
site adenosine's 2'OH reacts with phosphate group of nucleotide
at 5' end of intron (usually a "G") to form an ester linkage
between them.
C. Second transesterifcation reaction, also catalyzed by U2 and
U6 snRNAs: Oxygen of the now "free" 3'OH of the nucleotide at the
3' end of the upstream exon reacts with the 5' phosphate of the
nucleotide of the 5' end of the downstream exon forming another
ester linkage. Intron is released and exons are now spliced
together.
Since a given pre-mRNA can be spliced in alternative ways,
producing a variety of different mRNAs from the same pre-mRNA, with
each mRNA encoding a somewhat different polypeptide, this means
that a single gene can encode several different
polypeptides/proteins.
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Name _______________________________________________ 13 4.
Describe, in detail, the process of replication of DNA for creating
a lagging strand in prokaryotes. Be sure to name all the molecules
involved in the process and their functions.
1. DNA helicase unwinds DNA at the replication fork. 2.
Single-strand binding proteins (SSBs) bind to single-stranded DNA
template strand
to prevent formation of stem loops in that DNA strand
(intra-strand base pairing,) re-annealing of the two strands, or
binding of other nucleic acids.
3. As above occurs, (DNA) primase creates an RNA primer
complementary to a short stretch
of single-stranded template being produced by DNA helicase. 4.
Detachment of DNA polymerase from previous Okazaki fragment changes
conformation
of (Tau protein) clamp loader subunit of DNA polymerase
holoenzyme, inducing (gamma subunit of) clamp loader to load a
sliding clamp around DNA at 3' end of new RNA primer.
5. DNA polymerase re-localizes from previous Okazaki fragment to
3' end of new primer
and new clamp is attached to/loaded on DNA polymerase. Enzyme is
ready for synthesizing next Okazaki fragment
6. Starting at 3' end of an RNA primer made by (DNA) primase,
DNA polymerase reads
template strand 3 to 5, adding complementary nucleotides
one-by-one to new strand. Catalyzes reaction between 3 OH of last
nucleotide and -phosphate of incoming (next) nucleotide to create
an ester linkage (phosphodiester bond.) Creates Okazaki fragment
that lengthens in a direction opposite the movement of the
replication fork.
7. When DNA polymerase reaches 5' end of the RNA primer at the
5' end of previously
synthesized Okazaki fragment, DNA polymerase detaches from
template and moves to next primer (back to step 4.)
8. RNaseH digests most of RNA primer; exonuclease activity of
DNA polymerase (DNA pol I)
digests remainder of primer. 9. DNA polymerase fills in "gap"
created by digestion of RNA primer. 10. DNA ligase seals "nick" by
creating phosphodiester bond between 3' end of DNA that
replaced primer and 5' end of the previous Okazaki fragment.
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Name _______________________________________________ 14 5.
Assume a eukaryotic ribosome is in the middle of translation; i.e.
it has already synthesized part of the encoded polypeptide.
Describe one full cycle of translation, i.e. the events that occur
to add the next amino acid to the growing chain and to prepare the
ribosome for adding the next amino acid to the polypeptide. 1. A
new aminoacyl-tRNA, associated with EF-Tu-GTP, and with an
anticodon
complementary to mRNA codon now in small subunit of the A
(aminoacyl) site, enters the A-site of the ribosome.
2. The anticodon of this aminoacyl-tRNA base pairs with the mRNA
codon in small subunit portion of the A-site.
3. EF-Tu binding to A-site activates GTPase of EF-Tu,
hydrolyzing its GTP to GDP. 4. EF-Tu-GDP leaves ribosome and the
new aminoacyl-tRNA's 3'end with attached amino
acid is moved into peptidyl transferase center of the large
ribosomal subunit. 5. Peptidyl transferase (an activity of an rRNA
in the large subunit) catalyzes formation
of peptide bond between the amino group nitrogen of the amino
acid on the tRNA in the A-site and the terminal carbonyl group
carbon of the polypeptide attached to the tRNA in the P-site.
6. The "new" polypeptide (one amino acid longer) is transferred
to the "newest" tRNA residing in the A-site (though partially
displaced from the A site in the large ribosomal subunit, as it is
within the peptidyl transferase center.)
7. The elongation factor EF-G-GTP enters the large subunit
portion of the A site of the ribosome. Binding here brings EF-G-GTP
into factor binding center, which activates the GTPase of EF-G;
it's converted to EF-G-GDP causing a change in the conformation of
EF-G.
8. EF-G's change of conformation "pushes" tRNA in the A-site
completely from A-site into the P-site of ribosome; mRNA with codon
base-paired to this tRNA is "pulled" through ribosome one codon to
bring the next codon into A-site. Movement also "pushes" tRNA that
had been in the P-site to the E-site.
9. EF-G-GDP leaves ribosome, freeing A site for entry of next
aminoacyl-tRNA-EF-Tu complex.
10. tRNA in E-site leaves the ribosome.