Substrate preferences of O-methyltransferases in alfalfa suggest new pathways for 3-O-methylation of monolignols
Substrate preferences of O-methyltransferases in alfalfa suggest new pathways for 3-O-methylation of monolignols
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DOI：
10.1046/j.1365-313x.2001.00956.x
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发表时间：
2001-01-01
期刊：
影响因子：
7.2
通讯作者：
Dixon, RA
中科院分区：
文献类型：
--
作者：
Parvathi, K;Chen, F;Dixon, RA
在发育中的苜蓿茎提取物中，对单醇途径中所有潜在底物的相对0-甲基转移酶活性的测量显示活性顺序为：咖啡酰CoA >咖啡酰醇> 5-羟基阿魏酸>咖啡酰醛> 5-羟基松柏醇，5-羟基阿魏酰CoA > 5-羟基松柏醛>咖啡酸。所有活动的最大值发生在第七节间。在茎提取物与反义下调咖啡酰辅酶A O-甲基转移酶（CCoAOMT）的转基因苜蓿，活动与所有的基板，除了两个辅酶A酯不受影响。相反，咖啡酸O-甲基转移酶（COMT）的下调降低了对非代谢底物的活性，顺序为：5-羟基松柏醇> 5-羟基阿魏酸和咖啡醇>咖啡醛>咖啡酸> 5-羟基松柏醛。在大肠杆菌中表达的重组COMT对5-羟基松柏醛和咖啡醛的Vmax/Km值最高，对咖啡酸的Vmax/Km值最低。这些结果表明，COMT是不太可能甲基化咖啡酸在体内木质素生物合成过程中，并提供了一个替代途径，涉及咖啡酰醛和/或咖啡酰醇的COMT甲基化单木酚酶的证据。独立途径的概念，以愈创木基和乌灵基单木酚进行了讨论。
Measurement of relative O-methyltransferase activities against all potential substrates in the monolignol pathway in developing alfalfa stem extracts revealed activities in the order: caffeoyl CoA > caffeoyl alcohol > 5-hydroxyferulic acid > caffeoyl aldehyde > 5-hydroxyconiferyl alcohol, 5-hydroxyferuloyl CoA > 5-hydroxyconiferaldehyde > caffeic acid. Maxims for all activities occurred in the seventh internode. In stem extracts from transgenic alfalfa with antisense downregulated caffeoyl CoA O-methyltransferase (CCoAOMT), activities with all substrates except for the two coenzyme A esters were unaffected. In contrast, downregulation of caffeic acid O-methyltransferase (COMT) reduced activities against the non-esterifed substrates in the order: 5-hydroxyconiferyl alcohol > 5-hydroxyferulic acid and caffeoyl alcohol > caffeoyl aldehyde > caffeic acid > 5-hydroxyconiferaldehyde. Recombinant COMT expressed in Escherichia coli exhibited the highest V-max/K-m values with 5-hydroxyconiferaldehyde and caffeoyl aldehyde, and the lowest with caffeic acid. These results indicate that COMT is unlikely to methylate caffeic acid during lignin biosynthesis in vivo, and provide enzymatic evidence for an alternative pathway to monolignols involving methylation of caffeoyl aldehyde and/or caffeoyl alcohol by COMT. The concept of independent pathways to guaiacyl and syringyl monolignols is discussed.