Investigation of Tryptic Protein Digestion in Microdroplets and in Bulk Solution

• DOI: 10.1021/jasms.2c00072
• Publication Date: 2022-06-01T14:14:13Z
• Authors: Yongling Ai; Jeffrey Xu; Harsha P. Gunawar...; Richard N. Zare; Hao Chen
• Keywords: Deamidation; Digestion; Bottom-up proteomics

Recent studies have shown that ultrafast enzymatic digestion of proteins can be achieved in microdroplet within 250 μs. Further investigation peptides resulting from (MD) would necessary to evaluate it as an alternative the conventional bulk for bottom-up and biotherapeutic protein characterization. Herein we examined compared tryptic both MD solution. In case β-lactoglobulin B, preservation long was observed due short time. addition, is applicable digest high- low-abundance mixture. digesting NIST 8671 mAb antibody containing a low level commonly encountered host cell (HCP) PLBL2 (mAb:PLBL2 = 100:1 by weight), produced lower levels digestion-induced chemical modifications asparagine/glutamine deamidation, with overnight digestion. No significant difference between terms trypsin specificity based on examination semi- unspecific-cleaved peptides. Our study suggests MD, fast approach, could adopted proteomics research peptide mapping mAbs characterize site-specific deamidation glycosylation, purpose development biopharmaceuticals.