Purification and properties of a low-molecular-weight, high-alkaline pectate lyase from an alkaliphilic strain of Bacillus
Purification and properties of a low-molecular-weight, high-alkaline pectate lyase from an alkaliphilic strain of Bacillus
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DOI：
10.1271/bbb.63.65
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发表时间：
1999-01-01
影响因子：
1.6
通讯作者：
Ito, S
中科院分区：
文献类型：
--
作者：
Kobayashi, T;Koike, K;Ito, S
在芽孢杆菌属菌株KSM-P15的碱性培养物中发现低分子量、高碱性果胶酸裂解酶（果胶酸反式消除酶，EC 4.2.2.2），纯化至均一，并结晶。通过沉降平衡测定，该酶的相对分子量约为20，300，沉降系数（S-20，w（0））为1.73S。它是一种碱性蛋白质，等电点为pH 10.3，α-螺旋含量仅为6.6%。在Ca ~（2+）存在下，该酶以随机方式降解多聚半乳糖醛酸，生成4，5-不饱和寡聚半乳糖醛酸，在pH 10.5和50-55 ℃时具有最佳活性。它对棉纤维也有类似原果胶酶的活性。完整蛋白（28个氨基酸）和它的两个赖氨酰内肽酶切割的肽片段（8和12个氨基酸）的N-末端氨基酸序列与迄今报道的果胶酸裂解酶的序列相似性非常低。这些结果表明，芽孢杆菌菌株KSM-P15的果胶酸裂解酶可能是一种新的酶，属于一个新的家族。
A low-molecular-weight, high-alkaline pectate lyase (pectate transeliminase, EC 4.2.2.2) was found in an alkaline culture of Bacillus sp. strain KSM-P15, purified to homogeneity, and crystallized. The enzyme had a relative molecular weight of approximately 20,300 as measured by sedimentation equilibrium, with a sedimentation coefficient (S-20,w(0)) of 1.73S. It was a basic protein with an isoelectric point of pH 10.3, and the alpha-helical content was only 6.6%. In the presence of Ca2+ ions, the enzyme degraded polygalacturonic acid in a random manner to yield 4,5-unsaturated oligo-galacturonides and had its optimal activity around pH 10.5 and 50-55 degrees C. It also had a protopectinase-like activity on cotton fibers. The N-terminal amino acid sequences of the intact protein (28 amino acids) and its two lysyl endopeptidase-cleaved peptide fragments (8 and 12 amino acids) had very low sequence similarity with pectate lyases reported to date. These results strongly suggest that the pectate lyase of Bacillus sp. strain KSM-P15 may be a novel enzyme and belongs in a new family.