Indian Institute of Science Education and Research Bhopal

CHM 625: Biophysical Chemistry, Assignment 1

Posted on: 22/Aug/2017, submission date: 28/Aug/2017

You may prepare you assignment as a combination of results from a computer and hand written answers.
Members are expected to help each other within a group, but please do not copy answers from one
another. If copying is observed, all members of the group will get zero. The assignment would definitely
take several hours, so please work towards it until submission.

1. Towards the pursuit of determining primary sequencing of a protein, a researcher performed its
digestion (under reducing conditions) with enzymes pepsin and trypsin in two independent experiments.
The following peptide fragments for each of the assay was identified.
Pepsin: Pepsin preferentially cleaves at carboxylic end of aromatic amino acids and leucine.
Fragments EPPYTGPCKARIIRY, KSAEDCMRTCGGA, GGCRAKRNNF, NAKAGL, CQTF, RPDF,
FY, VY, CL. Trypsin: Trypsin preferentially cleaves at carboxylic end of positively charged amino acids.
Fragments RPDFCLEPPYTGPCK, AGLCQTFVYGGCR, SAEDCMR, YFYNAK, TCGGA, NNFK, IIR,
AK, AR, R. Mass spectroscopy revealed the molecular weight to be 6622 Da.
a. Determine the primary sequence of this protein using the above information.
b. Using the primary sequence that you have determined from the above question, perform a protein
BLAST search (https://blast.ncbi.nlm.nih.gov/Blast.cgi) and determine which protein this possibly could
be. Briefly describe the proteins function.
c. Download any high-resolution structure of this protein from the PDB and use any molecular
visualization software (like PyMol) to do the following.
d. Identify all secondary structure elements.
i. List the heavy atoms that are involved in H-bond formation across the protein along with their
distances (also report average and standard deviation).
ii. List the cysteine residues involved in disulfide bond formation. What is the average S-S bond
distance? Categorize them as right/left-handed disulfide bonds and list the value of dihedral angle.

2. 2008 chemistry Nobel Prize was awarded for the discovery and development of green fluorescent
protein to three eminent scientists: O. Shimomura, M. Chalfie and R.Y. Tsien. Why is a color associated
with this protein and how does this come up? Can this color be altered? How do you think this protein
can be used in Biophysical assays?

3. Genome related questions

a. What is the length of the human and common chimpanzee genome? List a few major differences that
arise between the genes that result in humans being different from the chimpanzee.
b. If you were to save the human genome sequence in a computer, how much space (in bytes) do you
think is required?

4. Provide the primary sequence of yeast tRNAPhe. List the modified residues present and their chemical
structure (only of nucleobase). Hint: RNA, 2000, 6, 1091-1105.

5. Each nucleosome consists of 147 bp of DNA wrapped around a compact octameric core containing
two molecules each of histones H2A, H2B, H3 and H4. Histones H2A/H2B and H3/H4 dimerize through
the non-covalent interaction of a highly conserved structural motif found in each core histone called the
histone fold, which also serves as the primary histoneDNA binding domain. - Nucleic Acids Res. 2002
Jan 1; 30(1): 341342.
Link: https://www.ncbi.nlm.nih.gov/research/HistoneDB2.0/, Database (2016). PMID: 26989147.
a. Use the above information to analyze the primary structure of the four histones, from this information
predict the portions that you anticipate to bind DNA.
b. Use the tertiary structure information to see how well your predicted agrees with your prediction from
the primary structure.
c. Use the primary and secondary structure information to pick amino acids that prefer to form -helices.

6. Suggest
(a) What content you would like to be added/changed in the course (individual answers expected)?
(b) As a group, but discussing only within your group, suggest a question that all of us could learn in the
field of biophysical chemistry.