The biosynthesis of
cholesterol is regulated mainly by
HMG-CoA reductase, however, recent studies indicated the pivotal role of another
enzyme in
cholesterol homeostasis. A previous report showed a marked decrease in the activity of
mevalonate pyrophosphate decarboxylase (MPD) in
stroke-prone spontaneously hypertensive rats and its possible involvement in the pathogenesis of the disorder. In this study, we purified liver MPD from rats fed a diet containing
cholestyramine and
pravastatin (CP diet) using conventional chromatographic techniques. We obtained two electrophoretically homogeneous
enzyme preparations; 45 and 37 kDa
proteins with specific activities of 8.0 and 7.4 micromol/min/mg, respectively. The
enzymes showed similar molecular weights of 90 kDa, as judged on gel permeation chromatography. A kinetic study indicated apparent Km values for
mevalonate pyrophosphate and
ATP of 22.7 microM and 0.71 mM, respectively, for the 45 kDa MPD, and 20.0 microM and 0.80 mM, respectively, for the 37 kDa MPD. Half maximum activities were observed at 1.5 mM and 1.1 mM Mg2+ for the 45 and 37 kDa MPDs, respectively. Both
enzymes required
ATP as a
phosphate acceptor, and in addition Mg2+, Mn2+, and Co2+ were effective as
divalent cations. The optimum pH for both
enzymes was 7.0. The isoelectric points for the 45 and 37 kDa MPDs were 5.6 and 5.4, respectively. Polyclonal antiserum raised against the 45 kDa
enzyme detected both the 45 and 37 kDa bands on immunoblots with CP diet-induced liver
crude extract as an
antigen. However, non-induced liver contained the 45 kDa
protein but not the 37 kDa
protein. These results indicated that the CP diet induced a new species, 37 kDa, of MPD which is characteristically and immunologically very similar to the well-known 45 kDa MPD.