Synthesis of high specific activity 35S-labelled N-methanesulfonyl farnesylcysteine and a photoactive analog

Tamara A. Kale, Conrad Raab, Nathan Yu, Evelyn Aquino, Dennis C. Dean, Mark D. Distefano

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8 Scopus citations

Abstract

Prenylated cysteine analogs, which mimic the prenylated cysteine residue of prenylated GTP-binding proteins (G-proteins), have been used in a variety of contexts for the study of prenylated G-protein behavior. In earlier work in this area, we prepared the photoactive analog [35S]4 and showed that it labelled RhoGDI upon photolysis; those results were consistent with the idea that GDI contains an isoprenoid binding site. Here, we describe the preparation of [35S]N-methanesulfonyl labelled analogs (1a and 2a) of N-acetyl farnesylcysteine and its methyl ester together with an improved synthetic procedure for photoactive analogs 3 and 4; specific activities of ∼1100 Ci/mmol were achieved. Compounds 1a and 2a in unlabelled form were used as competitors in photolysis reactions to show that the methanesulfonamido group is a reasonable acetamide substitution. Additional experiments show that the photoactive ester [35S]3 can cross-link GDI in both purified form and crude bacterial extract. However, the extent of cross-linking obtained with the ester ([35S]3) is significantly less than that observed with the free acid ([35S]4) despite the fact that the esterified form probably more closely reflects the structure of the C-terminus of a prenylated protein; using the GDI·Cdc42 co-crystal structure, the structural basis for these results is discussed.

Original languageEnglish (US)
Pages (from-to)29-54
Number of pages26
JournalJournal of Labelled Compounds and Radiopharmaceuticals
Volume46
Issue number1
DOIs
StatePublished - Jan 1 2003

Keywords

  • Isoprenoid binding site
  • Photoaffinity labelling
  • Prenylcysteine
  • RhoGDI
  • S

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