An improved method for preparing lysozyme with chemically 13C-Enriched methionine residues using 2-aminothiophenol as a reagent of thiolysis

Yoshito Abe, Tadashi Ueda, Taiji Imoto

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Jones et al. have reported that the ε-carbons of methionine residues in myoglobin can be enriched with stable isotope (13C) in two steps, i.e., methylation of methionine residues with 13CH3I in the protein and thiolysis using dithiothreitol. Using their method, we failed to prepare active lysozyme in which the ε-carbons of methionine residues are enriched with 13C, because many side reactions took place under the thiolysis condition (pH 10.5, 37°C). When we employed 2-aminothiophenol as a reagent for thiolysis, the reduction proceeded under a weakly acidic condition to afford fully active lysozyme, in which the ε-carbons of two methionine residues were enriched with 13C, in a 30% yield. Analysis of the 13C-edited NOESY spectra of 13C-enriched methionine lysozyme in the absence and presence of a substrate analogue indicated the occurrence of conformational change around Met 105 in lysozyme.

Original languageEnglish
Pages (from-to)1153-1159
Number of pages7
JournalJournal of Biochemistry
Volume122
Issue number6
DOIs
Publication statusPublished - Jan 1 1997

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology

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