Stereochemistry of valine and isoleucine biosynthesis

Hill, Richard K.; Sawada, Seiji; Arfin, Stuart M.

doi:10.1016/0045-2068(79)90003-8

Cited by 31 publications

(12 citation statements)

References 43 publications

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“…in the presence of the chiralshift reagent tris-[3-(heptafluoropropylhydroxymethylene)-(+)camphoratojeuropium(III), was shown to be greater than 99 0,h. Sodium salts were obtained by alkaline hydrolysis in nonepimerizing conditions (Hill et al, 1979;Crout et al, 1984) followed by the addition of HCI to bring the final pH of the substrate solutions to 8.2. For the synthesis of (2R)-and (2S)-2aceto-2-hydroxybutyrate, both ethyl esters were obtained by a reaction sequence similar to the preparation of (2R)-and (25)acetolactate ethyl esters, starting from either L-(+)or D-(-)lactic acid, after a stereoselective one-carbon homologation (Ortholand, 1991; a full description of the method is available from R. Douce on request).…”

Section: Protein Determinationmentioning

confidence: 99%

Isolation and kinetic properties of acetohydroxy acid isomeroreductase from spinach (Spinacia oleracea) chloroplasts overexpressed in Escherichia coli

Dumas

Job

Ortholand³

et al. 1992

Biochemical Journal

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Acetohydroxy acid isomeroreductase catalyses a two-step reaction, an alkyl migration and a NADPH-dependent reduction, in the assembly of the carbon skeletons of branched-chain amino acids. Detailed investigations of acetohydroxy acid isomeroreductase aimed at elucidating the biosynthetic pathway of branched-chain amino acids and at designing new inhibitors of the enzyme having herbicidal potency have so far been conducted with the enzymes isolated from bacteria. To gain more information on a plant system, the gene encoding the mature acetohydroxy acid isomeroreductase from spinach (Spinacia oleracea) leaf chloroplasts has been used to transform Escherichia coli cells and to overexpress the enzyme. A rapid protocol is described that allows the preparation of large quantities of pure spinach chloroplast acetohydroxy acid isomeroreductase. Kinetic and structural properties of the plant enzyme expressed in Escherichia coli are compared with those reported in our previous studies on the native enzymes purified from spinach chloroplasts and with those reported for the corresponding enzymes isolated from Escherichia coli and Salmonella typhimurium. Both the plant and the bacterial enzymes obey an ordered mechanism in which NADPH binds first, followed by substrate (either 2-acetolactate or 2-aceto-2-hydroxybutyrate). Inhibition studies employing an inactive substrate analogue, 2-hydroxy-2-methyl-3-oxopentanoate, showed, however, that the binding of 2-hydroxy-2-methyl-3-oxopentanoate and NADPH occurs randomly, suggestive of some flexibility of the plant enzyme active site. The observed preference of the enzyme for 2-aceto-2-hydroxybutyrate over 2-acetolactate is discussed with regard to the contribution of acetohydroxy acid isomeroreductase activity in the partitioning between isoleucine and valine biosyntheses. Moreover, the kinetic properties of the chloroplast enzyme support the notion that biosynthesis of branched-chain amino acids in plants is controlled by light. As judged by analytical-ultracentrifugation and gel-filtration analyses the overexpressed plant enzyme is a dimer of identical subunits.

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Section: Protein Determinationmentioning

confidence: 99%

Isolation and kinetic properties of acetohydroxy acid isomeroreductase from spinach (Spinacia oleracea) chloroplasts overexpressed in Escherichia coli

Dumas

Job

Ortholand³

et al. 1992

Biochemical Journal

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“…The reaction is likely to occur through the intermediacy of the corresponding phosphoric acid monoesters (R = PO 3 22 ). 4 Comparison with the mechanistically related steps involved in the biosynthesis of valine and isoleucine 11 (cf. Fig.…”

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confidence: 99%

Stereochemical course of the reduction step in the formation of 2-C-methylerythritol from the terpene precursor 1-deoxyxylulose in higher plants

Arigoni¹,

Giner²,

Sagner³

et al. 1999

Chem. Commun.

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“…This enzyme catalyzes an unusual two-step reaction: an alkyl migration in which the substrate, either 2-acetolactate (AL) or 2-aceto-2-hydroxybutyrate (AHB), is converted to 3-hydroxy-3-methyl-2-oxobutyrate or 3-hydroxy-3-methyl-2-oxopentanoate, respectively, followed by an NADPH-dependent reduction to give 2,3-dihydroxy-3isovalerate or 2,3-dihydroxy-3-methylvalerate, respectively (Figure 1). Studies from bacterial (Armstrong et al, 1974(Armstrong et al, , 1983Hill et al, 1979), fungi (Hawkes & Edwards, 1990), and plant enzymes (Dumas et al, 1992) have demonstrated that the 2S isomers of AL and AHB are the true substrates for the reaction, and that their diol products have respectively a configuration (2R) and (2R, 3R) (Hill & Yan, 1971;Crout & White-House, 1972). The reduction step requires the transfer of the pro-S hydrogen atom from NADPH (Arfin & Umbarger, 1969).…”

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confidence: 99%

Evidence for two catalytically different magnesium-binding sites in acetohydroxy acid isomeroreductase by site-directed mutagenesis

Dumas¹,

Butikofer²,

Job³

et al. 1995

Biochemistry

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Alignment of procaryotic and plant acetohydroxy acid isomeroreductase (EC 1.1.1.86) reveals five conserved regions designated domains I, II, III, IV, and V. Domain I has been previously proposed to correspond to the NADPH-binding site [Dumas et al. (1991) Biochem. J. 277, 469-475] and domain III to a putative magnesium-binding site [Sista & Bowman (1992) Gene 120, 115-118]. The binding and the function of this cation are of particular importance. First, Mg2+ is essential for the two-step reaction catalyzed by this enzyme: an isomerization followed by an NADPH-dependent reduction. Second, the plant acetohydroxy acid isomeroreductase exhibits Kd and Km values for Mg2+ of 5 microM and 6 microM, respectively. Such values correspond to the strongest affinity known between an enzyme and the metal ion. To determine if domain III of acetohydroxy acid isomeroreductase is effectively involved in magnesium binding, and with the goal to assign a function to the other conserved domains, site-directed mutagenesis was performed on each charged or polar conserved amino acids of domains II-V of the spinach acetohydroxy acid isomeroreductase. The results demonstrate that mutation of each of these amino acids leads to a partial or complete inactivation of enzyme activity. Steady-state kinetic analysis and equilibrium binding experiments show that both domains III and IV are directly involved in the binding of magnesium. Also, they suggest that magnesium bound to domain III plays a role in the reductive half-reaction, whereas, magnesium bound to domain IV is involved in the isomerization half-reaction.

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Stereochemistry of valine and isoleucine biosynthesis

Cited by 31 publications

References 43 publications

Isolation and kinetic properties of acetohydroxy acid isomeroreductase from spinach (Spinacia oleracea) chloroplasts overexpressed in Escherichia coli

Isolation and kinetic properties of acetohydroxy acid isomeroreductase from spinach (Spinacia oleracea) chloroplasts overexpressed in Escherichia coli

Stereochemical course of the reduction step in the formation of 2-C-methylerythritol from the terpene precursor 1-deoxyxylulose in higher plants

Evidence for two catalytically different magnesium-binding sites in acetohydroxy acid isomeroreductase by site-directed mutagenesis

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