Download Lab 4 Questions (Answers)

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LS1a Fall 2014
Lab 4 (PyMOL- Nucleic acids a proteins) question sheet
Q1) (6 points) The glycosidic bond links the 1’ carbon of deoxyribose with which atom of
adenine? Please be sure to include the number and name of the atom in your answer.
N9
Q2) (6 points) Which carbons of the two deoxyribose molecules are linked by the
phosphodiester bond? (Indicate the position of the carbons on the deoxyribose ring).
The 3’ carbon of one deoxyribose is linked to the 5’ carbon of the other deoxyribose by
the phosphodiester bond.
Q3) (6 points) Which nucleotide is at the 5’ end of this strand? Which nucleotide is immediately
3’ to that nucleotide?
The 5’ end is guanine (or “guanosine,” or “guanosine monophosphate”)
The base immediate 3’ to the 5’ terminal guanine is cytosine.
Q4) (6 points) Are the strands parallel or anti-parallel? Identify the 5’ and 3’ ends of each strand
to help you answer this question. Does the cyclic oxygen in the deoxyribose sugar point in
the same direction for both strands, or in different directions?
Antiparallel.
The cyclic oxygen in the deoxyribose moieties point in opposite directions on the two
antiparallel strands (or however the student chooses to convey the general point).
Q5) (6 points) Clearly label the major and minor grooves in the image below.
Minor
Major
Minor
Major
Major
Q6) (6 points) What are the distances between 1’ carbons of the sugars of an A-T base pair? How
does it relate to the distance between the 1’ carbons of the sugars of a G-C base pair? How
do these two distances contribute to the sequence-(in)dependence of the shape of the
double helix?
They are both 10.9 angtroms (Å). The fact that the 1’ carbons of deoxyribose are as far
apart in an A-T base pair as they are in a G-C base pair helps to allow a DNA double helix
to have the same shape regardless of its DNA sequence. The double helical structure and
geometry of DNA is sequence-independent.
Q7) (6 points) What is peculiar about the shape of the double-stranded DNA in this structure?
How does the electrostatic potential on the outside of CAP correspond with the
conformation of the DNA?
The DNA is severely bent around CAP rather than being straight (linear). The electrostatic
potential of CAP is mostly positive where it binds the DNA, and the positive charge of the
protein stabilizes the bend induced in the DNA.
Q8) (6 points) With what part of the DNA are the serine and threonine at the end of the
recognition helix interacting? Does this interaction convey information about the specific
DNA base sequence? What type of intermolecular interaction is this?
Serine and threonine are interacting with the phosphate group oxygens of the sugarphosphate backbone. These interactions do not convey sequence information about the
DNA bases. Since we do not know whether the phosphate group oxygens are charged or
not, these are either hydrogen bonds or dipole-ion interactions.
Q9) (15 points) Into what type of groove are the two arginines and the glutamate projecting
into? How do these three amino acid side chains specifically recognize the DNA sequence 5’TCxC-3’ on one strand (or 5’-GxGA-3’ on the complementary strand). The “x” stands for
bases that the protein is not specifically directly interacting with. Please be sure to include
specific intermolecular interactions in your answer.
One arginine is donating two hydrogen bonds (or is making two ion-dipole interactions)
with the 5’ guanine that his base-paired with the 3’ cytosine.
The glutamate is accepting a hydrogen bond (or making an ion-dipole interaction) with the
upstream cytosine as the other arginine is also donating two hydrogen bonds (or making
two ion-dipole interactions) with the guanine base-paired to the cytosine.
This same arginine is also hydrogen bonding (making an ion-dipole interaction) with the 5’
thymine (which is base-paired to the 3’ adenine on the other sequence).
5’ 3’
G C R donates two Hbonds to the two Hbond acceptors on G
x x No interactions are made
G C A different R donates two Hbonds to the two Hbond acceptors on G; E accepts an
Hbond from the Hbond donor on C
A T R donates a hydrogen bond to the Hbond acceptor on T
3’ 5’
[Because the crystal structures that we are looking at do not include hydrogens, it is
difficult to tell whether some interactions are hydrogen bonds or ion-dipole interactions.
It can be helpful to remind students to open Rich’s lecture 10 notes to look at the “Second
Genetic Code!” or for them to take out their section 5 handout to examine the major
groove and minor groove patterns.]
Q10) (6 points) How does the binding of cAMP help bring the two subunits of CAP (the orange
and the cyan subunits) together? What specific intermolecular interactions does cAMP
make with both CAP subunits?
cAMP binds CAP at the subunit interface, where it can form interactions with both
subunits. In particular, cAMP binds a serine on the cyan subunit and a cyan leucine places
its hydrophobic side chain near the nonpolar adenine ring of cAMP. The rest of the
interactions (a Thr, Ser, Arg, backbone NH, and Glu all hydrogen bond/ion-dipole) with
cAMP come from the orange subunit; an orange Val and Leu make van der Waals contacts
with cAMP.
Q11) (6 points) In addition to the polar interactions between the protein and cAMP, there are
also one orange valine, one orange leucine, and one cyan leucine shown. What part of cAMP
are they interacting with? What type of intermolecular interactions are they making with
cAMP?
The orange valine, orange leucine, and cyan leucine are making van der Waals interactions
with both hydrophobic faces of the adenine through van der Waals interactions (similar to
DNA base stacking).
Q12) (6 points) What are two interactions that are being made between the carboxy-terminal
arginine of CAP (in orange) with the alpha-CTD (in red)?
The CAP carboxy terminus is making an ionic bond with the side chain of one of the
arginines of the polymerase. The side chain of the CAP carboxy-terminal argineine is
donating hydrogen bonds (or making ion-dipole interactions) with a serine on the
polymerase.
Q13) (6 points) What is the difference between a DNA nucleoside and an RNA nucleoside? At
what carbon position is the difference located?
2’ hydroxyl
Q14) (6 points) What about the structure of this RNA molecule requires magnesium ions for
stability in the three locations where magnesium is indicated?
Since RNA is single stranded, as a single strand of RNA twists into it shape, it can compress
negative charges from its sugar-phosphate backbone near each other. These negative
charges would typically repel each other if they were not stabilized by magnesium ions.
Q15) (7 points) Draw the RNA-specific nitrogenous base “uracil” from the image of uracil
displayed in your PyMOL window. How is uracil chemically different than its DNA-specific
counterpart, thymine?
Uracil lacks a methyl group that thymine has.