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Pitfalls and Solutions in Mass Spectrometry-based Identification of Protein Glycation

ABSTRACT: Protein glycation is a type of post-translational modification (PTM) involving complex non-enzymatic reactions of reducing sugars or reactive dicarbonyls with proteins, generating a heterogeneous group of advanced glycation end-products (AGEs) such as carboxymethyllysine (CML). In bottom-up proteomics, 2-iodoacetamide (IAA) is the most commonly used reagent for cysteine alkylation before trypsin digestion, which exogenously gives a carbamidomethylation (CAM) group on the side chain of cysteine residue. However, offisite alkylation of IAA can occur at the protein N-terminus and other amino acid residues such as lysine residue. Here, for the first time, we provided evidence that IAA alkylation can result in false-positive identifications of CML, and N-isopropylacrylamide (NIPAM) can be used as an IAA alternative to avoid these pitfalls.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human) Escherichia Coli

TISSUE(S): Cell Culture

SUBMITTER: Humphrey, Wendong Ma

LAB HEAD: Terence Chuen Wai Poon

PROVIDER: PXD032235 | Pride | 2023-01-24

REPOSITORIES: Pride

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			Action	DRS
	20210115_HM_Ecoli_IAA_300minGr_R1.raw	Raw
	20210115_HM_Ecoli_IAA_300minGr_R2.raw	Raw
	20210115_HM_Ecoli_NIPAM_300minGr_R1.raw	Raw
	20210115_HM_Ecoli_NIPAM_300minGr_R2.raw	Raw
	20210115_HM_HEK_IAA_300minGr_R1.raw	Raw

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