Evaluation of the Effect of Trypsin Digestion Buffers on Artificial Deamidation

This work comprehensively compared 50 mM of ammonium acetate (pH 6), Tris-HCl (pH 8), ABC and TEAB as in-solution trypsin digestion buffers. Both iTRAQ quantification and label-free results indicate that ammonium acetate (pH 6) is more suitable than other buffers for studying endogenous deamidation and N-glycosylation due to the significant decrease of artificial Asn deamidation in comparison to other buffers without affecting protein and peptide identification and Gln deamidation. Determination of the half-life of Asn deamidation in the four buffers further validates the conclusion. Our results also indicate that among the commonly used tryspin digestion buffers, ABC and TEAB are not suitable for studying Asn deamidation and N-glycosylation, but Tris-HCl may be used if trypsin digestion has to be done at around pH 8.