Enzymic synthesis of lignin precursors. Further studies on cinnamyl-alcohol dehydrogenase from soybean-cell-suspension cultures.
Isoenzyme 2 of cinnamyl-alcohol dehydrogenase from soybean suspension cultures was purified about 3800-fold to apparent homogeneity by an improved purification procedure involving biospecific elution of the enzyme from a NADP+-agarose column. On sodium dodecylsulfate gels the dehydrogenase showed only one protein band with Mr 40 000 +/- 500. The enzyme is strongly inhibited by thiol reagents. Various metal chelators as well as the nonchelating 7,8-benzoquinoline also inhibited enzyme activity. Inhibition by 10 mM 1,10-phenanthroline could be partially reversed by addition of Zn2+. 1,10-Phenanthroline and 7,8-benzoquinoline are non-competitive inhibitors with respect to NADP+. The presence of zinc in the dehydrogenase was proved by atomic absorption spectroscopy and by specific incorporation of 65Zn into the enzyme. In steady-state kinetics inhibition patterns were obtained which are consistent with an ordered bi-bi mechanism in which NADP(H) is the first substrate to bind and the last product released. The cinnamyl-alcohol dehydrogenase belongs to the A-specific dehydrogenases and removes the pro-R hydrogen from coniferyl alcohol. The enzyme shows many similarities with alcohol dehydrogenases from horse and rat liver and from yeast.
D000429:Alcohol Oxidoreductases / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008031:Lignin / Q000096:biosynthesis*; D008099:Liver / Q000201:enzymology; D046911:Macromolecular Substances; D008970:Molecular Weight; D010944:Plants / Q000201:enzymology*; D020005:Propanols; D013025:Glycine max; D015032:Zinc
D007527:Isoenzymes; D046911:Macromolecular Substances; D020005:Propanols; D008031:Lignin; D000429:Alcohol Oxidoreductases; D015032:Zinc