Modification of Cysteine

Q1 Biochemistry, Genetics and Molecular Biology Current Protocols in Protein Science Pub Date : 2017-02-02 DOI:10.1002/cpps.22
Gregory A. Grant
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引用次数: 9

Abstract

This unit describes a number of methods for modifying cysteine residues of proteins and peptides. 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 described. Support protocols are included for recrystallization of iodoacetic acid, colorimetric detection of free sulfhydryls, and desalting of modified samples. © 2017 by John Wiley & Sons, Inc.

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半胱氨酸的修饰
本单元描述了一些修饰蛋白质和多肽的半胱氨酸残基的方法。提出了用卤代酰基试剂或n -乙基马来酰亚胺(NEM)对已知大小和组成的蛋白质中半胱氨酸残基进行烷基化的一般程序,并描述了在大小和组成未知和只有极少量蛋白质可用时使用的类似程序。还介绍了溴丙胺和N-(碘乙基)-三氟乙酰胺引入氨基的烷基化反应。描述了两种通常用于蛋白质的后续序列分析的程序,与4-乙烯基吡啶和丙烯酰胺烷基化,并且还介绍了吸附在测序膜上的4-乙烯基吡啶烷基化蛋白质的专门程序。介绍了半胱氨酸残基的亚砜水解可逆修饰,并提供了用甲酸氧化分析氨基酸组成的方案。描述了半胱氨酸残基暴露于空气中温和氧化为二硫化物。支持方案包括碘乙酸的重结晶,游离巯基的比色检测和改性样品的脱盐。©2017 by John Wiley &儿子,Inc。
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来源期刊
Current Protocols in Protein Science
Current Protocols in Protein Science Biochemistry, Genetics and Molecular Biology-Biochemistry
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期刊介绍: With the mapping of the human genome, more and more researchers are exploring protein structures and functions in living organisms. Current Protocols in Protein Science provides protein scientists, biochemists, molecular biologists, geneticists, and others with the first comprehensive suite of protocols for this growing field.
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Issue Information De Novo Protein Design Using the Blueprint Builder in Rosetta Methods for Expression of Recombinant Proteins Using a Pichia pastoris Cell-Free System Histone Purification Combined with High-Resolution Mass Spectrometry to Examine Histone Post-Translational Modifications and Histone Variants in Caenorhabditis elegans Issue Information
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