Involvement of the alpha subunit of farnesyl-protein transferase in substrate recognition.

Using photoaffinity labeling, we have identified a region in mammalian farnesyl-protein transferase (FPTase) involved in substrate recognition. The photolabel used (Compound 1) is a peptide containing the photoactive amino acid p-benzoylphenylalanine (Bpa). Upon exposure to UV light. Compound 1 inhibits FPTase activity in a time- and concentration-dependent manner. Photoinhibition of FPTase activity by Compound 1 is prevented by adding H-Ras to the reaction mixture, indicating that labeling is targeted to the enzyme active site. We used peptide mapping by HPLC, Edman sequencing, and matrix-assisted time-of-flight (MALDI-TOF) mass spectrometry to identify the site of interaction with radiolabeled Compound 1. These experiments indicate that a specific region of the alpha subunit of the enzyme, Asp110-Arg112, is involved in substrate binding and suggest that Glu111 is likely to be the residue covalently modified by the photoaffinity label. Sequence alignments between yeast and mammalian FPTases reveal that Glu111 is conserved. The implications of this finding are discussed in light of previous mutagenesis studies on FPTase.

Heterocyclic ureas: inhibitors of acyl-CoA:cholesterol O-acyltransferase as hypocholesterolemic agents.

A series of diaryl-substituted heterocyclic ureas was prepared, and their ability to inhibit acyl-CoA: cholesterol O-acyltransferase (ACAT) in vitro and to lower plasma total cholesterol in cholesterol-fed animal models in vivo was examined. N-(2,6-Diisopropylphenyl)-N'-tetrazole or isoxazole-substituted heterocyclic ureas proved optimal. A carbon chain of 11-14 carbons substituted 1,3 with respect to the amine provided the optimal side chain. Substitution of the alkyl chain generally lowered activity. Tetrazole urea 2i dosed at 3 mg/kg lowered plasma total cholesterol (TC) 67% in an acute, cholesterol-fed (C-fed) rat model of hypercholesterolemia and 47% in C-fed dogs. Tetrazole 2i, dosed at 10 mg/kg, also lowered TC 52% and raised HDL cholesterol 113% in rats with pre-established hypercholesterolemia.

Antiatherosclerotic activity of inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase in cholesterol-fed rabbits: a biochemical and morphological evaluation.

Atherosclerotic lesion development was assessed in the thoracic aorta and chronically denuded iliac-femoral artery of hypercholesterolemic New Zealand White rabbits using inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase which have previously been shown to possess varying degrees of hepatoselectivity in rats. Atorvastatin, previously known as CI-981 (2.5 mg/kg), PD135022 (1.0 mg/kg), simvastatin (2.5 mg/kg), lovastatin (2.5 mg/kg), PD134965 (1.0 mg/kg), pravastatin (2.5 mg/kg) and BMY22089 (2.5 mg/kg) were added to a 0.5% cholesterol, 3% peanut, 3% coconut oil diet and fed for 8 weeks. Although reductions in plasma total cholesterol of 27% to 60%, VLDL-cholesterol of 31% to 71% and plasma total cholesterol exposure of 37% to 43% were obtained, no correlation between these parameters and vascular lipid content, lesion size or monocyte-macrophage content was noted. Iliac-femoral lipid content was unchanged; however, atorvastatin and simvastatin significantly reduced the cholesterol content of the thoracic aorta by 45%-62%. Atorvastatin and PD135022 reduced the size of the iliac-femoral lesion by 67% and monocyte-macrophage content by 72%. Simvastatin, lovastatin and PD134965 decreased the monocyte-macrophage content; however, lesion size was unchanged. Pravastatin and BMY22089 had no effect on lesion size or content. No compound significantly reduced the extent of thoracic aortic lesions. We concluded that changes in plasma lipids and lipoproteins noted with the various HMG-CoA reductase inhibitors did not account for the beneficial effect on atherosclerotic lesion development. The antiatherosclerotic potential of the HMG-CoA reductase inhibitors was compound-specific and clearly not a class effect.

Analysis of commercial samples of dimethyl sulfoxide.

Refractive index measurements and spectral data (IR and NMR) were used to verify the purity claimed by commercial samples of dimethyl sulfoxide.