CHEM 251

Exam #3 12/02/13
1. (25 pts)
a. (10 pts) Draw the chemical mechanism for charging a tRNA with an amino acid.

b. (10 pts) Describe the roles of EF-Tu and EF-G in catalyzing the elongation cycle
during protein synthesis. Your answer should include consideration of the various sites of
tRNA binding, and the involvement, if any, of guanosine nucleotides.
EF-Tu: forms ternary complex with aminoacyl-tRNA : EF-Tu.GTP.aa-tRNA.
Cognate ternary complex binds rapidly to A-site of ribosome in response to codon
GTP is hydrolyzed, EF-Tu.GDP dissociates from ribosome, leaving aa-tRNA bound
to ribosome where it cam participate in peptide bond formation.
EF-G.GTP complex binds to pretranslocation complex and catalyzes movement of
peptidyl-tRNA and deacylated tRNA from the A and P sites to the P and E sites,
respectively. This translocation occurs following GTP hydrolysis.

c. (5 pts) tRNA synthetases must charge specific tRNAs. Indicate and name three regions
on a generic cloverleaf structure of tRNA that are used by various tRNA synthetases to
distinguish one tRNA from another.
4 regions - anticodon, 3-terminus, variable loop, 3(acceptor)-stem

Variable
loop

2. (25 pts 5 each)

Briefly describe the following, using figures as appropriate
a. Lariat formation during splicing of eukaryotic RNA.

OH
5'-

5'-exon

-O-PO2O-

intron

-O-PO2O-

3'-exon

-3'

O
5'-

5'-exon

-O-H

O2PO-

intron

-O-PO2O-

3'-exon

-3'

b. Poly(A) Binding Protein

Binds the poly (A) tail on the 3 end of a eukaryote. It helps to protect the 3 end
from exonuclease action during the time it takes mRNA to migrate from the
nucleus to the cytoplasm.
c. linc RNA
long intergenic noncoding RNA that regulate gene expression - LincRNAs are
transcribed by RNA polymerase II. Most lincRNAs are processed similarly to
protein-coding RNAs, including 5-end capping, 3- end polyadenylation, splicing
of introns, and intra- cellular transport.
d. IRES
Internal Ribosome Entry Site Allows cap-independent initiation
of protein synthesis. The IRES is a highly folded structure in the 5UTR of mostly viral mRNAs that binds to the 40S ribosomal small
subunit.

e. Shine-Dalgarno helix
At the translation initiation step, a polypurine sequence 3-8 (or several) nucleotides
upstream from initiator codon (or to the 5-side) base pairs with a polypyrimidine
stretch at the 3-terminus of 16S rRNA to form the SD helix. An SD helix can also
form during polypeptide elongation when there are polypurine sequences upstream
of the P-site codon.

3. (20 pts)
a. (5 pts) How would you design a tyrosyl-tRNA synthetase so that it efficiently
discriminated between tyrosine and phenylalanine as a substrate?
Aminoacyladenylate formation site large enough to accommodate Tyr (and
therefore Phe), with favorable hydrophobic interactions to bind aromatic ring and a
favorable H-bonding interaction with the Tyr OH.
Editing (aminoacyladenylate hydrolysis) site with favorable hydrophobic
interactions to bind aromatic ring but too small to accommodate OH group
b. (5 pts) Draw the structure of a glycerophospholipid that has two fatty acyl groups (16:1
cis and an 18:2 trans, trans) and a phosphorylserine head group.

R1 =

CH3(CH2)6

R2 =
CH3(CH2)6

X = CH2CH(NH3+)CO2-

(CH2)6

(CH2)6

c. (10 pts)
I. Glycogen phosphorylase b is an allosteric dimeric enzyme having two
conformations, an inactive form (T state) and an active form (R state), with the T
state favored in the absence of ligands. The substrate, inorganic phosphate, Pi,
binds preferentially to the R state. ATP is an allosteric inhibitor and AMP is an
allosteric activator. Draw curves of enzyme-catalyzed velocity vs. [Pi]:
i.
in the absence of allosteric effectors;
ii.
in the presence of ATP
iii.
in the presence of AMP
+AMP

Phosphorylase
activity

No addition

+ATP

[Inorganic phosphate]

II. Rationalize the effects of ATP and AMP from an overall metabolic perspective.
Active glycogen phosphorylase catalyzes glycogen breakdown, ultimately leading to
ATP formation. High [ATP] acts as a feedback inhibitor. High AMP, which
correlates with low [ATP], is stimulatory

4. (30 pts)
Write chemical structures and mechanisms (arrow pushing) for the following enzymecatalyzed reactions. Note that some or all of these reactions might i) have more than one
step and ii) utilize a cofactor. When cofactors are present, you need only include portions
of the structure that are chemically relevant.
a. (12 pts)
OH
2-O PO-CH CHCHO + Pi + NAD
3
2

HO O
||
2-O PO-CH CHCOPO 2- + NADH
3
2
3

b. (12 pts)

CH2OH
CHO

CH2OH
H

C=O
HO
H

H
OH
CH2OPO32-

H
H

C=O
CHO

OH
OH
OH
CH2OPO32-

HO
H

OH

OH

OH
CH2OPO32-

OH
CH2OPO32-

c. (6 pts)

OPO3-PO32-

OPO3-PO32-

+ PPi