Goodwin G W, Rougraff P M, Davis E J, Harris R A
Department of Biochemistry, Indiana University School of Medicine, Indianapolis 46202-5122.
J Biol Chem. 1989 Sep 5;264(25):14965-71.
Methylmalonate semialdehyde dehydrogenase was purified from rat liver in order to define the distal portion of valine catabolism and related pathways in mammals. The purified enzyme is active with malonate semialdehyde and consumes both stereoisomers of methylmalonate semialdehyde, implicating a single semialdehyde dehydrogenase in the catabolism of valine, thymine, and compounds catabolized by way of beta-alanine. The oxidation of malonate and methylmalonate semialdehydes by this enzyme is CoA-dependent, the products being acetyl-CoA and propionyl-CoA, respectively. Expected activity with ethylmalonate semialdehyde as substrate was not found. Methylmalonate semialdehyde dehydrogenase was separated on DEAE-Sephacel into two isoforms which differ in mobility during nondenaturing polyacrylamide gel electrophoresis. The two forms are immunologically cross-reactive and exhibit the same N-terminal sequence, suggesting that one form is the product of the other. The monomer molecular mass, determined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, was 58 kDa. The native molecular mass, estimated by gel filtration, was 250 kDa, suggesting a tetrameric structure.
为了确定缬氨酸分解代谢的远端部分以及哺乳动物中的相关途径，从大鼠肝脏中纯化了甲基丙二酸半醛脱氢酶。纯化后的酶对丙二酸半醛有活性，且能消耗甲基丙二酸半醛的两种立体异构体，这表明在缬氨酸、胸腺嘧啶以及通过β-丙氨酸分解代谢的化合物的分解过程中涉及单一的半醛脱氢酶。该酶对丙二酸和甲基丙二酸半醛的氧化作用依赖于辅酶A，产物分别为乙酰辅酶A和丙酰辅酶A。未发现以乙基丙二酸半醛为底物时的预期活性。甲基丙二酸半醛脱氢酶在DEAE-葡聚糖凝胶上分离为两种同工型，在非变性聚丙烯酰胺凝胶电泳中迁移率不同。这两种形式具有免疫交叉反应性，且显示相同的N端序列，表明一种形式是另一种形式的产物。在十二烷基硫酸钠存在下通过聚丙烯酰胺凝胶电泳测定的单体分子量为58 kDa。通过凝胶过滤估计的天然分子量为250 kDa，表明其为四聚体结构。
