Question

6. a. If the oligopeptide AFVRWGTHLESKLVMSLDIHLRF were cleaved completely by reaction with trypsin and V8 protease and the products subjected to ESI TOF-MS in positive ion mode with the peptides in a pH 2 solution, which product will reach the detector last? Explain why. b. If the Lys on the oligopeptide were to be acetylated on its side chain, would the last peptide to reach the detector be changed? Explain. c. To determine the M, of a purified protein, you subject it to ESI TOF-MS in positive ion mode with the protein in a pH 2 solution. The spectrum shows a series of peaks with the two most prominent neighboring peaks having m/z ratios of 648.78 and 659.52. What is the uncharged mass of the protein (i.e., M, at its pl) in the 648.78 peak? d. Is the charge on the protein in part c consistent with the protein having a typical basic amino acid composition? Justify your conclusion mathematically.

Answer 1

The product which reach detector last is Lys and Arg. The net charge of ESI TOF MS is positive ion, in reaction with trypsin and V8 protease it contains a carboxylic acid group, the a-amino group, and a second amino group attached to the side chain are deprotonated first. Carboxyl side of basic Amino Acids Lys & Arg is the end product .In a very acidic solution, pH=2, all of these groups are protonated. Therefore the overall net charge of the peptide is positive, the peptide is considered basic.