Peptide N-terminal Bead Capture and Release

ABSTRACT: The field of proteomics has expanded recently with more sensitive techniques for the bulk measurement of peptides as well as single-molecule techniques. One limiting factor for these methods is the need for multiple chemical derivatizations and highly pure proteins free of contaminants. We show a solid-phase capture strategy suitable for the proteolysis, purification, and subsequent chemical modification of peptides. We use this resin on an HEK293T cell lysate and perform one-pot proteolysis, capture, and derivatization to generate a cellular proteome that identified over 8,000 proteins. We also show that this capture can be reversed in a traceless manner, such that it is amenable for single-molecule proteomics techniques. With this technique, we perform a fluorescent labeling and C-terminal differentiation on a peptide and subject it to fluorosequencing demonstrating that washing the resin is sufficient to remove excess dyes prior to single-molecule protein sequencing.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Embryonic Stem Cell, Kidney

SUBMITTER: Brendan Floyd  

LAB HEAD: Edward Marcotte

PROVIDER: PXD016291 | Pride | 2021-06-01

REPOSITORIES: Pride