Publications

• Structural and biochemical characterization of the Bacillus cereus 3-hydroxyisobutyrate dehydrogenase.

  Park S.C., Kim P.H., Lee G.S., Kang S.G., Ko H.J., Yoon S.I.

  The 3-hydroxyisobutyrate dehydrogenase (HIBADH) family catalyzes the NAD(+)- or NADP(+)-dependent oxidation of various β-hydroxyacid substrates into their cognate semialdehydes for diverse metabolic pathways. Because HIBADH group members exhibit different substrate specificities, the substrate-rec ... >> More

  The 3-hydroxyisobutyrate dehydrogenase (HIBADH) family catalyzes the NAD(+)- or NADP(+)-dependent oxidation of various β-hydroxyacid substrates into their cognate semialdehydes for diverse metabolic pathways. Because HIBADH group members exhibit different substrate specificities, the substrate-recognition mode of each enzyme should be individually characterized. In the current study, we report the biochemical and structural analysis of a HIBADH group enzyme from Bacillus cereus (bcHIBADH). bcHIBADH mediates a dehydrogenation reaction on S-3-hydroxyisobutyrate substrate with high catalytic efficiency in an NAD(+)-dependent manner; it also oxidizes l-serine and 3-hydroxypropionate with lower activity. bcHIBADH consists of two domains and is further assembled into a functional dimer rather than a tetramer that has been commonly observed in other prokaryotic HIBADH group members. In the bcHIBADH structure, the interdomain cleft forms a putative active site and simultaneously accommodates both an NAD(+) cofactor and a substrate mimic. Our structure-based comparative analysis highlights structural motifs that are important in the cofactor and substrate recognition of the HIBADH group. << Less

  Biochem Biophys Res Commun 474:522-527(2016) [PubMed] [EuropePMC]

• Purification and characterization of 3-hydroxyisobutyrate dehydrogenase from rabbit liver.

  Rougraff P.M., Paxton R., Kuntz M.J., Crabb D.W., Harris R.A.

  3-Hydroxyisobutyrate dehydrogenase (3-hydroxy-2-methyl propanoate: NAD+ oxidoreductase, EC 1.1.1.31) was purified 1800-fold from rabbit liver by detergent extraction, differential solubility in polyethylene glycol and (NH4)2SO4, and column chromatography on DEAE-Sephacel, phenyl-Sepharose, CM(carb ... >> More

  3-Hydroxyisobutyrate dehydrogenase (3-hydroxy-2-methyl propanoate: NAD+ oxidoreductase, EC 1.1.1.31) was purified 1800-fold from rabbit liver by detergent extraction, differential solubility in polyethylene glycol and (NH4)2SO4, and column chromatography on DEAE-Sephacel, phenyl-Sepharose, CM(carboxymethyl)-Sepharose, Affi-Gel Blue, and Ultrogel AcA-34. The enzyme had a native Mr of 74,000 and appeared to be a homodimer with subunit Mr = 34,000. The enzyme was specific for NAD+. It oxidized both S-3-hydroxyisobutyrate and R-3-hydroxyisobutyrate, but the kcat/Km was approximately 350-fold higher for the S-isomer. Steady state kinetic analysis indicates an ordered Bi Bi reaction mechanism with NAD+ binding before 3-hydroxyisobutyrate. The enzyme catalyzed oxidation of S-3-hydroxyisobutyrate between pH 7.0 and 11.5 with optimal activity between pH 9.0 and 11.0. The enzyme apparently does not have a metal ion requirement. Essential sulfhydryl groups may be present at both the 3-hydroxyisobutyrate and NAD+ binding sites since inhibition by sulfhydryl-binding agents was differentially blocked by each substrate. The enzyme is highly sensitive to product inhibition by NADH which may play an important physiological role in regulating the complete oxidation of valine beyond the formation of 3-hydroxyisobutyrate. << Less

  J Biol Chem 263:327-331(1988) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Structure, interactions and action of <i>Mycobacterium tuberculosis</i> 3-hydroxyisobutyric acid dehydrogenase.

  Srikalaivani R., Singh A., Vijayan M., Surolia A.

  Biochemical and crystallographic studies on <i>Mycobacterium tuberculosis</i> 3-hydroxyisobutyric acid dehydrogenase (<i>Mt</i>HIBADH), a member of the 3-hydroxyacid dehydrogenase superfamily, have been carried out. Gel filtration and blue native PAGE of <i>Mt</i>HIBADH show that the enzyme is a d ... >> More

  Biochemical and crystallographic studies on <i>Mycobacterium tuberculosis</i> 3-hydroxyisobutyric acid dehydrogenase (<i>Mt</i>HIBADH), a member of the 3-hydroxyacid dehydrogenase superfamily, have been carried out. Gel filtration and blue native PAGE of <i>Mt</i>HIBADH show that the enzyme is a dimer. The enzyme preferentially uses NAD⁺ as the cofactor and is specific to <i>S</i>-hydroxyisobutyric acid (HIBA). It can also use <i>R</i>-HIBA, l-serine and 3-hydroxypropanoic acid (3-HP) as substrates, but with much less efficiency. The pH optimum for activity is ∼11. Structures of the native enzyme, the holoenzyme, binary complexes with NAD⁺, <i>S</i>-HIBA, <i>R</i>-HIBA, l-serine and 3-HP and ternary complexes involving the substrates and NAD⁺ have been determined. None of the already known structures of HIBADH contain a substrate molecule at the binding site. The structures reported here provide for the first time, among other things, a clear indication of the location and interactions of the substrates at the active site. They also define the entrance of the substrates to the active site region. The structures provide information on the role of specific residues at the active site and the entrance. The results obtained from crystal structures are consistent with solution studies including mutational analysis. They lead to the proposal of a plausible mechanism of the action of the enzyme. << Less

  Biochem J 475:2457-2471(2018) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.