Enzymatic properties and deduced amino acid sequence of a high-alkaline pectate lyase from an alkaliphilic Bacillus isolate
Enzymatic properties and deduced amino acid sequence of a high-alkaline pectate lyase from an alkaliphilic Bacillus isolate
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DOI：
10.1016/s0304-4165(99)00017-3
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发表时间：
1999-04-19
影响因子：
3
通讯作者：
Ito, S
中科院分区：
文献类型：
--
作者：
Kobayashi, T;Hatada, Y;Ito, S
研究了一种高碱性果胶酸裂解酶（pectate trans-eliminase，EC 4.2.2.2.）从嗜碱芽孢杆菌属菌株KSM-P7（命名为Pel-7）纯化至均一。经SDS-聚丙烯酰胺凝胶电泳测定，纯化的Pel-7分子量约为33 kDa。等电点接近或高于pH 10.5。在Ca 2+离子存在下，Pel-7以随机方式反式消除聚半乳糖醛酸以产生寡聚半乳糖醛酸;其在pH 10.5和大约60至65 ℃的甘氨酸-NaOH缓冲液中表现出最佳活性。Mn 2+和Sr 2+离子可以在几乎与Ca 2+离子相同的水平上充当辅因子。它还表现出原果胶酶样活性，从棉纤维中释放可溶性果胶和/或寡聚半乳糖醛酸。对pel基因进行了克隆和序列测定，推测的成熟Pel-7的氨基酸序列（302个氨基酸，33，355个Dal）与已知的Pel家族的氨基酸序列同源性为27 ~ 32%。此外，Pel-7似乎具有与其他Pel相似的平行β-螺旋和活性位点拓扑结构的核心结构，如通过Pel蛋白的二级结构预测所揭示的。这些结果表明，Pel-7基本上属于Pel超家族，虽然酶和分子性质不同。(C)1999 Elsevier Science B. V.保留所有权利。
A high-alkaline pectate lyase (pectate trans-eliminase, EC 4.2.2.2.) from alkaliphilic Bacillus sp. strain KSM-P7, designated Pel-7, was purified to homogeneity. The purified Pel-7 had a molecular mass of approximately 33 kDa as determined by SDS-polyacrylamide gel electrophoresis. The isoelectric point was close to or higher than pH 10.5. In the presence of Ca2+ ions, Pel-7 trans-eliminated polygalacturonate in random manner to generate oligogalacturonides; it exhibited optimal activity at pH 10.5 and around at 60 to 65 degrees C in glycine-NaOH buffer. Mn2+ and Sr2+ ions can serve as cofactors at almost the same level of Ca2+ ions. It also exhibited a protopectinase-like activity, liberating soluble pectin and/or oligogalacturonides from cotton fibers. The pel gene was cloned and sequenced, and the deduced amino acid sequence of mature Pel-7 (302 amino acids, 33, 355 Dal showed some conserved regions in Pel superfamily, although homology to amino acid sequences of known Pels with 27 to 32% identity. Furthermore, Pel-7 appears to have similar core structure of parallel beta-helix and active site topology with other Pels as revealed by secondary structure prediction in the Pel proteins. These results suggest that Pel-7 is basically grouped into Pel superfamily although the enzymatic and molecular properties are different. (C) 1999 Elsevier Science B.V. All rights reserved.