1. the first doubt is the discrepancy in experimental and theoretical pI/Mr, the protein candidates usually have a different value in 1-2 unit between experimental and theoretical pI, and 10-20 kDa in molecular weight (sometimes double!). I have considered that the difference in pI may due to modification (such as phosphorylation) and difference in Mr may due to dimerization or native protein structure. But I didnít have strong evidence to support my thought.
2. and Iím planning to confirm the expression of these protein candidates using western blotting on SDS-PAGE. So, which Mr is the true one expected to appear on the blot? Will the experimental one or theoretical one appear on the blot?
3. Iíve heard that we could get the idea of the function of the unknown or hypothetical protein from bioinformatics. Do you have any suggestion and experience about that?
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peptide mass fingerprinting doubts
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