Download File

Name:_____________________________
Student ID:______________________
Exam I
Summer 2012
Some Facts that you may find helpful:
Typical pKa’s
Terminal a-carboxyl group = 3.1
Aspartic acid and Glutamic acid = 4.1
Histidine = 6.0
Terminal a-amino group = 8.0
Tyrosine = 10.9
Lysine = 10.8
Arginine = 12.5
1. Indirect ELISA is used to detect _________________.
a. Antigen
b. Antibody
c. DNA
d. Virus
2. The pKa of lactic acid is 3.86. What is the pH of a buffer in which the concentration of lactic
acid is 0.2M and concentration of sodium lactate is 2.0M
a. 2.86
b. 3.86
c. 4.86
d. 7.00
3. Which of the following is true of the isoelectric point of amino acids?
a. The isoelectric points of all amino acids are the same
b. It is the pH at which an amino acid conducts electricity
c. It is the pH at which an amino acid has equal numbers of positive and negative charges
d. All of the above are true
a
c
b
4. In the picture above what does a and b represent respectively?
a. Beta sheet, left-handed helix
b. Right-handed helix, beta sheet
c. Right-handed helix, left-handed helix
d. Beta sheet, right-handed helix
5. Watson-Crick base pairs are held together by
a. Van der Waals forces
b. Hydrogen bonds
c. Covalent bonds
d. Hydrophobic attraction
6. Which of the following statements about the peptide bond are true?
a. The peptide bond is planar because of the partial double-bond character of the bond
between the carbonyl carbon and the nitrogen.
b. There is relative freedom of rotation of the bond between the carbonyl carbon and the
nitrogen.
c. The hydrogen that is bonded to the nitrogen atom is trans to the oxygen of the carbonyl
group.
d. There is no freedom of rotation around the bond between the  carbon and the carbonyl
carbon.
7. You have 2L of solution of HCl that has a pH of 3.4. What is the number of moles of HCl needed
to make this solution? (4 points)
8. Answer the following questions about the peptide HRCE.
a. Draw the structure of the peptide at physiological pH. (8 points)
b. Label a peptide bond, -carbon, N-terminus, C-terminus. (4 points)
c. What would be the net charge of the peptide at pH 3.5 and pH 11.5. (4 points)
9. When complimentary sequences of DNA are mixed in solution they will anneal to form double
stranded helices.
a. What is the second law of thermodynamics? (2 points)
The total entropy (disorder) of a system plus that of its surrounding always increases
b. Why does this appear to disobey the second law of thermodynamics? (2 points)
This appears to disobey the second law because when two strands of DNA anneal, this decreases
disorder (increases order)
c. Explain why it does not disobey the second law of thermodynamics. (4 points)
This does not disobey the second law of thermodynamics because when two strands of DNA anneal
energy is released to the surrounding to cause a universal increase in disorder.
10. Explain five differences between a  helix found in proteins and a helix found in the B form of
DNA. (Be specific) (5 points)
There are many differences. A few examples are:  helix is made up of amino acids while B form is
made up of nucleotides,  helix is single stranded while B form is double stranded,  helix has 3.6 amino
acids per turn while B form has 10 nucleotides per turn,  helix is found in protein while B form is found
in DNA,  helix has H-bond with the peptide back bone while B form has hydrogen bonds between bases
and also has vanderwall and hydrophobic interactions to stabilize the secondary structure,  helix is the
same throughout whild B form has major and minor groove.
11. Answer the following question regarding the DNA sequence:
5’-ATGCAGCCGCTT-3’
a. Determine the mRNA sequence coded in the DNA. (2 points)
b. Determine the amino acid sequence coded by the DNA. (2 points)
c. If the DNA sequence had a mutation that changed it to ATGCAACCGCTT, explain why
or why not this would have a deleterious effect on the protein? (3 points)
There would be no change because the mutation still encodes for the same amino acid.
12. What is tertiary structure? Describe five types of interactions that are important for the formation
of tertiary structure? Give an example of an amino acid that is able to do each type of interaction.
(12 points)
Tertiary structure- defines the specific overall 3-D shape of the protein
Types of interactions that are important for the formation
1.
Hydrogen bondattractive interaction between a hydrogen atom and an electronegative atom, such as nitrogen,
oxygen or fluorine (ex. YRDESTHKWRNQ)
2.
Vander walls - The
distribution of electric charge around an atom changes with time
at any instant, charge distribution is asymmetric. encourages similar asymmetry around
neighboring atoms leading to attraction (ex. Any amino acid)
3.
Hydrophobic - nonpolar
molecules show an increased tendency to associate with one another (ex. VLIPGAFYWM)
4.
Electrostatic – interaction
between opposite charges (ex. RKEDH)
Metal Ion bridge – two
5.
ions with the same charge are linked together by a metal ion (RKEDH)
Disulfide bond –
6.
covalent bond between two C residue
13. Below is a restriction map of a plasmid DNA. Complete the agarose gel on the right by drawing
bands of appropriate size in the lanes under the names of the restriction endonucleases that were
used to digest the plasmid. (6 points)
EcoRI, BamHI
BamHI, SalI
SalI
EcoRI
1.2 kb
BamHI
55
7 kb
SalI
2.6kb
3.0 kb
SalI
14. Answer the following question in regards to the picture below. (12 points)
a. What type of bond is represented by a? b-N-glycosidic bond
b. What type of bond is represented by b? phosphodiester bond
c. What is the name of the base represented by c? quanine
d. What is the name of the base represented by d? adenine
e. What is the name of the base represented by e? thymine
f. Is the base represented by f a purine or pyrimidine? pyrimidine
g. Label the 5’ and 3’ ends of both chains.
h. Is this structure DNA or RNA, explain your answer?
DNA because it is made up of deoxyribonucleic acid (deoxyribose sugar) and there is a thymine.
i. Why is it important for so that the purine to form Watson and Crick interactions with a
pyrimidine? It is important that the distance between the two strands remains constant.
15. A protein is overexpressed by recombinant DNA technology. The protein is expressed in the
cytoplasm of E. coli. The protein is composed of a peptide made up of 200 amino acids, with a
theoretical pI of 4.5. It is known that this protein has a quaternary structure composed of at least
two of these peptide chains.
a. Describe how you would purify this protein to approximately 90% purity. (4points)
There are many answers that would be correct. But first you must break open (lyse the cells). This could
be done by sonication, homogenization, detergent, freeze thaw…etc). Then you would do crude
fractionation (centrifuge to remove the insoluble portion) followed by a purification step. Examples
would be ion exchange chromatography, salting out, gel filtration, dialysis, ….etc. You would also
require a way to tract the purification (ex. SDS-PAGE, enzyme activity, 2D gel electrophoresis, …..etc.
b. Describe a method that you could use to determine how many peptide chains make up the
quaternary structure of the protein. (4points)
Compare the molecular weight of the native protein compared to the molecular weight of the denature
protein. (example, use gel filtration to determine the size of the native protein then compare it to the size
from SDS_PAGE. For example the gel filtration gave a size of 40kD and the SDS-PAGE gave a size of
20kD that means two polypeptide chains make up the quaternary structure.
c. Describe how you could determine if disulfide bonds are involved in formation of the
quaternary structure. (3 points)
To determine if disulfide bonds are involved you would use urea to denature the protein
and determine the size. Then you would use urea plus a reducing agent such as BME
or DTT to break the disulfide bonds and determine the size of the protein. If disulfide
bonds were involved in forming the quaternary structure then you would only get the
size of the 200 amino acid peptide chain when using the reducing agent.