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

Abstract

AbstractPrenylated 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. Copyright © 2002 John Wiley & Sons, Ltd.