Synthesis of the protein cutting reagent iron (S)-1-(p- bromoacetamidobenzyl)ethylenediaminetetraacetate and conjugation to cysteine side chains

Douglas P. Greiner, Reiko Miyake, Justin K. Moran, A. Daniel Jones, Tomofumi Negishi, Akira Ishihama, Claude F. Meares

Research output: Contribution to journalArticlepeer-review

56 Scopus citations

Abstract

Convenient methodology for preparation and conjugation of the protein- cutting iron chelate iron (S)-1-(p- bromoacetamidobenzyl)ethylenediaminetetraacetate (Fe-BABE) is given. This formulation of the reagent can be handled in a manner analogous to many other protein-labeling reagents, such as fluorescent probes or cross-linkers. By taking advantage of the recently discovered peptide hydrolysis reaction, the chelate may be tethered to a single site (e.g., a cysteine side chain) and used to map its proximity to individual peptide bonds by automated Edman sequencing of the protein fragments produced. The method is illustrated by conjugation of Fe-BABE to the carboxy terminal domain (amino acid residues 234-329) of the Escherichia coli RNA polymerase alpha subunit. The molecular mass of the protein conjugate was confirmed by electrospray ionization mass spectrometry.

Original languageEnglish (US)
Pages (from-to)44-48
Number of pages5
JournalBioconjugate Chemistry
Volume8
Issue number1
DOIs
StatePublished - Jan 1997

ASJC Scopus subject areas

  • Chemistry(all)
  • Organic Chemistry
  • Clinical Biochemistry
  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry

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