Photoreactive Analogues of Prenyl Diphosphates as Inhibitors and Probes of Human Protein Farnesyltransferase and Geranylgeranyltransferase Type I

Photoreactive analogues of prenyl diphosphates have been useful in studying prenyltransferases. The effectiveness of analogues with different chain lengths as probes of recombinant human protein prenyltransferases is established here. A putative geranylgeranyl diphosphate analogue, 2-diazo-3,3,3-trifluoropropionyloxy-farnesyl diphosphate (DATFP-FPP), was the best inhibitor of both protein farnesyltransferase (PFT) and protein geranylgeranyltransferase-I (PGGT-I). Shorter photoreactive isoprenyl diphosphate analogues with geranyl and dimethylallyl moieties and the DATFP-derivative of farnesyl monophosphate were much poorer inhibitors. DATFP-FPP was a competitive inhibitor of both PFT and PGGT-I with Ki values of 100 and 18 nM, respectively. [32P]DATFP-FPP specifically photoradiolabeled the β-subunits of both PFT and PGGT-I. Photoradiolabeling of PGGT-I was inhibited more effectively by geranylgeranyl diphosphate than farnesyl diphosphate, whereas photoradiolabeling of PFT was inhibited better by farnesyl diphosphate than geranylgeranyl diphosphate. These results lead to the conclusions that DATFP-FPP is an effective probe of the prenyl diphosphate binding domains of PFT and PGGT-I. Furthermore, the β-subunits of protein prenyltransferases must contribute significantly to the recognition and binding of the isoprenoid substrate.