Unit

UNIT 15.1 Modification of Cysteine

  1. Mark W. Crankshaw,
  2. Gregory A. Grant

Published Online: 1 MAY 2001

DOI: 10.1002/0471140864.ps1501s03

Current Protocols in Protein Science

Current Protocols in Protein Science

How to Cite

Crankshaw, M. W. and Grant, G. A. 2001. Modification of Cysteine. Current Protocols in Protein Science. 3:15.1:15.1.1–15.1.18.

Author Information

  1. Washington University School of Medicine, St. Louis, Missouri

Publication History

  1. Published Online: 1 MAY 2001
  2. Published Print: MAR 1996

Abstract

This unit describes a number of methods for modifying cysteine residues of proteins and peptides by reduction and alkylation procedures. A general procedure for alkylation of cysteine residues in a protein of known size and composition with haloacyl reagents or N-ethylmaleimide (NEM) is presented, and alternate protocols describe similar procedures for use when the size and composition are not known and when only very small amounts of protein are available. Alkylations that introduce amino groups using bromopropylamine and N-(iodoethyl)-trifluoroacetamide are also presented. Two procedures that are often used for subsequent sequence analysis of the protein, alkylation with 4-vinylpyridine and acrylamide, are described, and a specialized procedure for 4-vinylpyridine alkylation of protein that has been adsorbed onto a sequencing membrane is also presented. Reversible modification of cysteine residues by way of sulfitolysis is described, and a protocol for oxidation with performic acid for amino acid compositional analysis is also provided. Gentle oxidation of cysteine residues to disulfides by exposure to air is detailed. Support protocols are included for recrystallization of iodoacetic acid, colorimetric detection of free sulfhydryls, and desalting of modified samples.