    |
||||
|
xxx |
H. Peter Spielmann Professor B.S., University of California, Los Angeles, 1984 Ph.D., University of California, Berkeley, 1991 Post-doctoral Fellow, University of California, Berkeley, 1991-95 NIH Post-doctoral Fellow, 1992-94 hps@pop.uky.edu 859-257-4790
 Farnesylation is a post-translational lipid modification in which a 15-carbon farnesyl isoprenoid is linked via a thioether bond to specific cysteine residues of proteins in a reaction catalyzed by protein farnesyltransferase (FTase). We synthesized the benzyloxyisoprenyl pyrophosphate (BnPP) series of transferable farnesyl pyrophosphate (FPP) analogs (1a-e) to test the length dependence of the isoprenoid substrate on the FTase catalyzed transfer of lipid to protein substrate. Kinetic analyses show that pyrophosphates 1a-e and geranyl pyrophosphate (GPP) transfer with a lower efficiency than FPP whereas geranylgeranyl pyrophosphate (GGPP) does not transfer at all. While a correlation was found between Km and analog hydrophobicity and length, there was no correlation between kcat and these properties. Potential binding geometries of FPP, GPP, GGPP and analogs 1a-e were examined by modeling the molecules into the active site of the FTase crystal structure. We found that analog 1d displaces approximately the same volume of the active site as does FPP, where GPP and analogs 1a-c occupy lesser volumes and 1e occupies a slightly larger volume. Modeling also indicated that GGPP adopts a different conformation than the farnesyl chain of FPP, partially occluding the space occupied by the Ca1a2X peptide in the ternary x-ray crystal structure. Within the confines of the FTase pocket, the double bonds and branched methyl groups of the geranylgeranyl chain significantly restricts the number of possible conformations relative to the more flexible lipid chain of analogs 1a-e. The modeling results also provide a molecular explanation for the observation that an aromatic ring is a good isostere for the terminal isoprene of FPP.
|
||
