This is a highly advanced Biochemistry question a. Retrieve FASTA (sequence) abd

Question

This is a highly advanced Biochemistry question

a. Retrieve FASTA (sequence) abd PDB files for human (5ikp) and rabbit (2gj4) glycogen phosphorylase (GP) from the PDB database.

b. Display the position of AMP binding on a ribbon diagram of human GP, then align the primary (FASTA) structures by CLUSTALW, and show the region in sequence where AMP binds to human/rabbit GP.

Tips

Google or search for the PDB files on the PDB website. These sites should have all of the information about the proteins that you need to complete the rest (Download Files->PDB and FASTA). The PDB files should then be opened with Chimera. Find the residue(s) to which AMP (make sure you are looking at AMP) binds by hovering over the shown residues. Remember these residues for the next part! Send me a picture of the AMP binding site.

Using this website (or any other alignment site) http://www.ebi.ac.uk/Tools/msa/clustalo/ (Links to an external site.) , you should put the FASTA sequences in here. Then, find the residues you remembered from the last part. Take a picture and highlight or just tell me what the residues are so I can see them when you send me the aligned sequences.

Explanation / Answer

Answer:

a. 5IKP: Fasta sequence as obtained from the fasta sequece file in the PDB:

2GJ4: Fasta sequence as obtained from the fasta sequece file in the PDB:

b. The residues involved in AMP binding to human glycogen phosphorylase can be easily determined using Pymol, UCSF Chimera or PISA.

The residues involved in AMP binding are: (PDB: 5IKP)

AMP (number 902 in the pdb file) interacts with Tyr196 (Chain A, distance=3.11 Angstroms), Arg309 (ChainA, distance=2.79 Angstroms) through hydrogen bonds. There are 34 non bonded interactions out of which the most potent are Gln71 and Tyr75.

The sequence alignment file for 5IKP and 2GJ4 is given below:

The hydrogen bonding residues Y196 and R309 (in the 4th and 6th line of the sequence alignment) have been marked (Bold, italics and underlined).

The corresponding residues in 2GJ4 are F196 (Phe196) and R309 and have been marked (Bold, italics and underlined)

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