Mevalonate Kinase Activity Research Articles

3-Hydroxy-3-methylglutaryl CoA reductase and mevalonate kinase of Neurospora crassa

Two enzymes of polyisoprenoid synthesis, 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase (mevalonate:NADP oxidoreductase [acylating CoA], EC 1.1.1.34) and mevalonate kinase (ATP:mevalonate 5-phosphotransferase, EC 2.7.1.36), are present in the microsomal and soluble fractions of Neurospora crassa, respectively. HMG CoA reductase specifically uses NADPH as reductant and has a K(m) for dl-HMG CoA of 30 micro M. The activities of HMG CoA reductase and mevalonate kinase are low in conidia and increase threefold during the first 12 hr of stationary growth. Maximum specific activities of both enzymes occur when aerial hyphae and conidia first appear (2 days), but total activities peak later (3-4 days). Addition to the growth media of ergosterol or beta-carotene, alone or in combination, does not affect the specific or total activity of either enzyme. The mevalonate kinase of N. crassa, purified 200-fold to a specific activity of 5 micro moles/min/mg, is free from HMG CoA reductase, phosphomevalonate kinase, ATPase, adenylate kinase, and NADH oxidase activities. Mevalonate kinase specifically requires ATP as cosubstrate and exhibits a marked preference for Mg(2+) over Mn(2+), especially at high ratios of divalent metal ion to ATP. Kinase activity is inhibited by p-hydroxymercuribenzoate, and this inhibition is partially prevented by mevalonate or MgATP. Optimum activity occurs at pH 8.0-8.5 and at about 55 degrees C. The Neurospora kinase, like that of hog liver, has a sequential mechanism for substrate addition. The Michaelis constants obtained were 2.8 mM for dl-mevalonate and 1.8 mM for MgATP(-2). Geranyl pyrophosphate is an inhibitor competitive with MgATP (K(i) = 0.11 mM).

Intrachloroplastidic mevalonate-activating enzymes in plant cells

Mevalonate phosphorylation by chloroplasts from Spinacia oleracea leaves, Agave americana leaves and Pinus pinaster seedlings is reported. Different modifications in the technique of chloroplast isolation by using an aqueous method have been introduced in order to protect the mevalonate kinase and phosphomevalonate kinase activities. The acid pH values of the preparations obtained by conventional methods from Agave americana and Pinus pinaster decrease their kinase activities; therefore, 0.35 M TrisHCl buffer, pH 7.9, containing 0.35 M mercaptoethanol must be used as the isolation medium. Ultrasonic treatment has been found to be the best method for chloroplast disruption in Spinacia oleracea and Agave americana, but a considerable inactivation of mevalonate kinase from Pinus pinaster has been observed after this treatment. The time course of mevalonic acid (MVA) phosphorylation by intrachloroplastidic enzymes was studied. Preparations from etiolated leaves of Agave americana also phosphorylated MVA, but their plastids are more fragile than those from green leaves.

Isolation of two fractions with mevalonate kinase activity from Pinus pinaster and Agave americana

Isolation of two fractions with mevalonate kinase activity from Pinus pinaster and Agave americana

Dietary sterol effects on mevalonate kinase activity in axenic and non-axenic Sarcophaga bullata (Diptera)

The effects of dietary sterols on the mevalonate kinase of Sarcophaga bullata larvae have been determined. Larvae reared axenically to day 6 on semi-defined cholesterol and β-sitosterol diets evidence a two- to threefold increase in enzyme specific activity compared to the non-axenic larvae at the same developmental time. Under non-axenic growth conditions, the larva evidences a clear pattern of kinase activity in response to varying dietary cholesterol concentrations. A 50 per cent increase in specific activity is observed when cholesterol concentration is reduced to 1 100 that of the standard dietary optimum (1·0 per cent). Of other sterols studied, β-cholestanol produces the most significant effect on the larval enzyme. Larval growth rate indices and survival determinations have been calculated for the insects reared on cholesterol and β-sitosterol regimens. The results of these investigations are discussed in relation to the effect of nutritional stimuli on enzyme levels in axenic and non-axenic organisms and the control of isoprenoid biosynthesis in this insect.

The activity and kinetic properties of mevalonate kinase in superovulated rat ovary.

Methods are described for the assay and partial purification of mevalonate kinase from superovulated rat ovary. The total activity of mevalonate kinase in superovulated rat ovary was 1.6+/-0.14units/g wet wt.; it was unchanged by the administration of luteinizing hormone in vivo. The K(m) of a partially purified preparation of mevalonate kinase for dl-Mevalonate was 3.6+/-0.5mum; its K(m) for MgATP(2-) was 120+/-7.7mum. The enzyme was inhibited by geranyl pyrophosphate and farnesyl pyrophosphate, but not by isopentenyl pyrophosphate or 3,3'-dimethylallyl pyrophosphate. dl-mevalonate 5-phosphate inhibited at high concentrations. With both geranyl pyrophosphate and farnesyl pyrophosphate the inhibition was competitive with respect to MgATP(2-). The K(i) for inhibition by geranyl pyrophosphate was 1.3+/-0.2mum; the K(i) for inhibition by farnesyl pyrophosphate was 1.0+/-0.3mum. These findings are discussed with reference to the control by luteinizing hormone of steroidogenesis from acetate.