Investigation of the action patterns of pectinmethylesterase isoforms through kinetic analyses and NMR spectroscopy - Implications in cell wall expansion
Investigation of the action patterns of pectinmethylesterase isoforms through kinetic analyses and NMR spectroscopy - Implications in cell wall expansion
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DOI：
10.1074/jbc.273.50.33150
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发表时间：
1998-12-11
影响因子：
4.8
通讯作者：
Goldberg, R
中科院分区：
文献类型：
--
作者：
Catoire, L;Pierron, M;Goldberg, R
将特征良好的果胶样品与来自绿豆下胚轴的果胶甲酯酶的细胞壁结合和溶解的纯异构体一起孵育在pH5.6和pH7.6条件下，沿脱酯途径每隔沿着测定酶活性和平均产物结构。并通过（13）C MMR和电位测量来探测酯化度。与β同工酶相比，在α和γ同工酶的行为中观察到二分法。理想的嵌段脱酯化机理再现了整个反应过程中的实验平均结构（甲酯分布）。在α和γ亚型的情况下，可以假设在pH 5.6下与第二近邻位置处的游离羧基相关的单链机制，而在pH 7.6下需要一些多重攻击特征来再现数据。与前述机制不同的几种机制与两种pH值下β亚型的数据一致。无论是在这些反应中产生的多糖的性质和果胶甲酯酶在细胞壁硬化过程中的作用沿着生长梯度进行了讨论。
Well characterized pectin samples were incubated with cell wall-bound and -solubilized pure isoforms of pectinmethylesterase from mung bean hypocotyls (Vigna radiata), Both enzyme activity and average product structure were determined at intervals along the deesterification pathway at pH 5.6 and 7,6, The latter analyses were performed by (13)C MMR spectroscopy, and the degree of esterification was probed by both (13)C MMR and potentiometric measurements. A dichotomy was observed in the behavior of the alpha and gamma isoforms when compared with that of the beta isoenzyme. Ideal blockwise deesterification mechanisms reproduced the experimental average structures (methylester distribution) throughout the course of the reaction. In the case of the alpha and gamma isoforms, a single chain mechanism associated with a free carboxyl group at the second nearest neighbor position could be postulated at pH 5.6, whereas some multiple attack character was required to reproduce the data at pH 7.6. Several mechanisms that differed from the preceding ones were compatible with the data for the beta isoform at the two pH values. Both the nature of the polysaccharides produced in these reactions and the role of pectinmethylesterase in the cell wall-stiffening process along the growth gradient are discussed.