Photoaffinity Heterobifunctional Cross-Linking Reagents Based on N-(Azidobenzoyl) tyrosines

Nobuyuki Imai, Tadashi Kometani, Peter J. Crocker, Jean B. Bowdan, Ayhan Demir, Lori D. Dwyer, Dennis M. Mann, Thomas C. Vanaman, David S. Watt

Research output: Contribution to journalArticlepeer-review

9 Scopus citations

Abstract

New heterobifunctional cross-linking reagents that possessed a photoactive terminus, an electrophilic terminus, and a linking arm between the two termini that had a radiolabeled, enzymatically cleavable bond were synthesized. In a model study, succinimidyl N-[N′-(4-azidobenzoyl)tyrosyl]-β-alanate (16A) was coupled to n-butylamine (a Lys surrogate), iodinated, and cleaved with chymotrypsin in the presence of tyrosylamide to afford the desired adduct N-(N′-(4-azidobenzoyl)-3-iodotyrosyl)tyrosinamide, thereby demonstrating the feasibility of the enzymatic cleavage. In a biochemical study, succinimidyl N-[N′-(3-azido-5-nitrobenzoyl)tyrosyl]-β-alanate (16C) was coupled to Lys-75 of calmodulin (CaM), and the radioiodinated monoadduct was successfully photo-cross-linked, in a calciumdependent manner, to the human erythrocyte plasma membrane Ca2+,Mg2+-ATPase and to a synthetic fragment (M13) containing the CaM-binding region of myosin light-chain kinase. In the latter case, densitometry readings indicated 20% cross-linking efficiency.

Original languageEnglish
Pages (from-to)138-143
Number of pages6
JournalBioconjugate Chemistry
Volume1
Issue number2
DOIs
StatePublished - Mar 1 1990

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Biomedical Engineering
  • Pharmacology
  • Pharmaceutical Science
  • Organic Chemistry

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