Question

Two vials containing the A- and β-chains of human insulin mysteriously lost their labels. Assuming no reference standard could be found, how can one determine which vial contains which chain? Please propose all experiments you would use, the expected results in sufficient details for each experiment, and your rationales

Answer 1

1) the best experiment is to perform the SDS PAGE gel run.

the A chain is of 21 amino acid and B chain is of 30 amino acid so both the chain will migrate different distance in 15% or 17.5% SDS PAGE reducing gel.

on the gel, the band close to the well is B chain and the band far from well is of A chain.

2) we can also perform N terminal amino acid sequencing.

the first amino acid of A chain is Glycine and first amino acid of B chain is Phenylalanine.

3) we can also perform amino acid analysis of both sample-

A chain contains 4 Cys while B chain contains 2 Cys residue. this will show a double peak intensity shift of Cys in A compared to B chain amino acid analysis. ( intensity of Cys residue is more in A as compared to b chain)

4) we can also perform MALDI analysis of both sample. the sample having high Mw is B chain and having less Mw is A chain.