title: Enzymic synthesis of lignin precursors. Comparison of cinnamoyl-CoA reductase and cinnamyl alcohol:NADP+ dehydrogenase from spruce (Picea abies L.) and soybean (Glycine max L.).
abstract: Cambial sap of spruce (Picea abies) proved to be a good source for isolation of cinnamoyl-CoA reductase and cinnamyl alcohol:NADP+ dehydrogenase. Apparently homogeneous enzymes were obtained by a multistep procedure including dye-ligand chromatography and for the reductase also affinity chromatography on (coenzyme A)-agarose. An improved purification procedure for the reductase from soybean cell cultures is also reported. Molecular weights and subunit composition of reductase and dehydrogenase from spruce are very similar to those of the corresponding enzymes from soybean. Reduction of feruloyl-CoA to coniferaldehyde catalysed by the reductase is a freely reversible reaction with an equilibrium constant of 5.6 x 10(-4) M at pH 6.25. A strong dependence of the Michaelis constants on the type of buffer was found. For reductase the Km-value of feruloyl-CoA in phosphate buffer (5.2 microM) is about 14-times similar than in citrate buffer (73 microM). Pronounced differences in substrate specificities between the enzymes from spruce and soybean were found, which reflect the different lignin composition of gymnosperms and dicotyledenous angiosperms. From the kinetic constants of the enzymes it can be concluded that under physiological conditions feruloyl-CoA is the preferred substrate for the reductase from both sources whereas sinapoyl-CoA is a substrate only for the soybean reductase and sinapyldehyde a substrate only for the soybean dehydrogenase. 4-Coumaroyl-CoA is a poor substrate for the reductase from both spruce and soybean. This result is consistent with the low content of 4-coumaryl alcohol units in gymnosperm and angiosperm lignin.
mesh_terms: D000429:Alcohol Oxidoreductases / Q000302:isolation & purification*; D000445:Aldehyde Oxidoreductases / Q000302:isolation & purification*; D055598:Chemical Phenomena; D002621:Chemistry; D004792:Enzyme Precursors / Q000138:chemical synthesis*; D008031:Lignin / Q000096:biosynthesis*; D010944:Plants / Q000201:enzymology*; D013025:Glycine max / Q000201:enzymology*; D013045:Species Specificity; D013379:Substrate Specificity
chemical_list: D004792:Enzyme Precursors; D008031:Lignin; D000429:Alcohol Oxidoreductases; C018656:cinnamyl alcohol dehydrogenase; D000445:Aldehyde Oxidoreductases; C019904:cinnamoyl CoA reductase