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In vivo, enzymatic reduction of some protein disulfide bonds, allosteric disulfide bonds, provides an important level of structural and functional regulation. The free cysteine residues generated can be labeled by maleimide reagents, including biotin derivatives, allowing the reduced protein to be detected or purified. During the screening of monoclonal antibodies for those specific for the reduced forms of proteins, we isolated OX133, a unique antibody that recognizes polypeptide resident, N-ethylmaleimide (NEM)-modified cysteine residues in a sequence-independent manner. OX133 offers an alternative to biotin-maleimide reagents for labeling reduced/alkylated antigens and capturing reduced/alkylated proteins with the advantage that NEM-modified proteins are more easily detected in mass spectrometry, and may be more easily recovered than is the case following capture with biotin based reagents.

Original publication

DOI

10.1080/19420862.2016.1152443

Type

Journal article

Journal

MAbs

Publication Date

05/2016

Volume

8

Pages

672 - 677

Keywords

Allosteric, detection, disulfide bond, labile, maleimide, reduction, Animals, Antibodies, Monoclonal, Cysteine, Ethylmaleimide, HIV Envelope Protein gp120, Oxidation-Reduction, Protein Processing, Post-Translational