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

10 Scopus citations


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
Issue number2
StatePublished - Mar 1 1990

Bibliographical note

Funding Information:
We thank the NationalScience Foundation (Grant CHE- 8607441) fortheir financial support, K. Drauz andA. Bernd ofDegussaAG for agenerous gift ofamino acids, NATO for a travel grant (Grant RG #0346/88) to D.S.W. and A.D.,andtheUniversityofKentuckyforthepurchase ofbond issueequipmentandforFacultyGrantInitia- tiveAwards (toD.S.W.andT.C.V.).

ASJC Scopus subject areas

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


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