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Question

Download Krizek et al. A concensus Zinc Finger Peptide...' J. Amer. Chem. Soc. 1991, vol 113, pp 4518 (a) Discuss the method they used for creating a consensus peptide. (b) In 1991, there were 131 zinc finger sequences known, which came from 18 proteins. How many sequences are there now? How many complete 3 dimensional structures? (c) The first structure determined for a zinc finger is PDB ID IZNF. Concentrating only on the secondary structure (select-cartoon in PYMOL) draw a contact map of this molecule (by hand). (d) The authors state that protonated histidines were not observed to bind metal ions - binding to neutral histidines is an either/or process, in the absence of metal ions, the two histidines ligands each have pKa's of 6.5. What is the binding energy for these two protons together? How do they compare with the binding energies for Zn^2+ and Co^2+ (calculate these as well)? How were each of these binding constants determined? If you determined the histidine pKa's in the presence of a small concentration, say 10 mu M, of metal ion, which of the two ions would cause the effective pKa to be lowest? How could you experimentally determine the entropic and enthalpic contributions to binding either protons or metal ions to this protein? Which do you think would have the higher entropic cost, protonation or metal binding? Why? (e) While the first known zinc finger proteins bind double-stranded DNA, later a class of these proteins were found to bind to RNA molecules at the double, stranded part of a stem-loop. What is a stem-loop? Make up an RNA sequence that you think will form a stem loop structure. Label which part(s) will form the stem and which will form the loop.

Explanation / Answer

4 (a) Krizek et al., has adapted the method to create a consensus Zinc finger peptide based on the overall shape of the protein backbone in the folded domain with the use of a data base of 131 zing finger sequence. The fold group consider by Krizek et al., are the Cys2His2 in the domain. Cys2His2 form a knuckle and two more form the c terminus of a helix. such type of consensus zinc finger peptide is best characterized class of zinc fingers and are very common in mammalian transcription factors.

b. 131, None

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