Covalent modification of stathmin by CCNU determined by FTMS analysis of modified proteins and tryptic peptides.

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Wu WW, Wang G, Liang XJ, Park JK, Shen RF

Covalent modification of stathmin by CCNU determined by FTMS analysis of modified proteins and tryptic peptides.

Biochem Biophys Res Commun. 2008 Feb 29;367(1):7-13. Epub 2007 Dec 26.

PubMed ID

18162179 [ View in PubMed

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Abstract

Chemical modification of proteins is often carried out to generate protein-small molecule conjugates for various applications. The high resolution and mass accuracy of a Fourier transform mass spectrometer is particularly useful for assessing the extent or sites of covalent modifications. As protein-small molecule reactions often produce products with variable numbers of the compound incorporated at different sites, a direct mass analysis of the reaction products at times yields mass spectra hard to interpret. Chromatographic separation at protein level could reduce the complexity of a sample, thus allowing more accurate mass spectrometric analysis. In this report, we demonstrate the utility of reversed-phase protein chromatography and FT-ICR mass spectrometry in analyzing CCNU (lomustine, 1-(2-chloroethyl)-3-cyclohexyl-1-nitroso-urea, MW: 233.7Da) modification of stathmin. With this combined approach, we determined the stoichiometry as well as sites of CCNU incorporation into the protein, demonstrating differential reactivity of several lysyl residues to CCNU alkylation.

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Drug Targets

Drug	Target	Kind	Organism	Pharmacological Action	Actions
Lomustine	Stathmin-4	Protein	Humans	Unknown	Antagonist	Details