"Investigation of Tryptic Protein Digestion in Microdroplets and in Bul" by Yongling Ai, Jeffrey Xu et al.
 

Investigation of Tryptic Protein Digestion in Microdroplets and in Bulk Solution

Document Type

Article

Publication Date

7-6-2022

Abstract

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

Identifier

85131967758 (Scopus)

Publication Title

Journal of the American Society for Mass Spectrometry

External Full Text Location

https://doi.org/10.1021/jasms.2c00072

e-ISSN

18791123

ISSN

10440305

PubMed ID

35647885

First Page

1238

Last Page

1249

Issue

7

Volume

33

Grant

1R15GM137311-01

Fund Ref

National Institutes of Health

This document is currently not available here.

Share

COinS