Department of Food Science and Biotechnology, Kyungsung University, Busan 608-736, Korea.
Appl Microbiol Biotechnol. 2012 Sep;95(6):1643-53. doi: 10.1007/s00253-012-4266-y. Epub 2012 Jul 18.
A gene for a polyMG-specific alginate lyase possessing a novel structure was identified and cloned from Stenotrophomas maltophilia KJ-2 by using PCR with homologous nucleotide sequences-based primers. The recombinant alginate lyase consisting of 475 amino acids was purified on Ni-Sepharose column and exhibited the highest activity at pH 8 and 40 °C. Interestingly, the recombinant alginate lyase was expected to have a similar catalytic active site of chondroitin B lyase but did not show chondroitin lyase activity. In the test of substrate specificity, the recombinant alginate lyase preferentially degraded the glycosidic bond of polyMG-block than polyM-block and polyG-block. The chemical structures of the degraded alginate oligosaccharides were elucidated to have mannuronate (M) at the reducing end on the basis of NMR analysis, supporting that KJ-2 polyMG-specific alginate lyase preferably degraded the glycosidic bond in M-G linkage than that in G-M linkage. The KJ-2 polyMG-specific alginate lyase can be used in combination with other alginate lyases for a synergistic saccharification of alginate.
从嗜麦芽寡养单胞菌 KJ-2 中，通过使用基于同源核苷酸序列的引物的 PCR，鉴定并克隆了一个编码具有新颖结构的多聚 MG 特异性褐藻胶裂解酶的基因。由 475 个氨基酸组成的重组褐藻胶裂解酶在 Ni-Sepharose 柱上进行纯化，并在 pH8 和 40°C 时表现出最高的活性。有趣的是，重组褐藻胶裂解酶预计具有类似软骨素 B 裂解酶的催化活性位点，但没有表现出软骨素裂解酶活性。在底物特异性测试中，重组褐藻胶裂解酶优先降解聚 MG 嵌段的糖苷键，而不是聚 M 嵌段和聚 G 嵌段。基于 NMR 分析，降解的褐藻胶寡糖的化学结构被阐明在还原端具有甘露糖酸盐 (M)，支持 KJ-2 多聚 MG 特异性褐藻胶裂解酶优先降解 M-G 键中的糖苷键，而不是 G-M 键中的糖苷键。KJ-2 多聚 MG 特异性褐藻胶裂解酶可与其他褐藻胶裂解酶联合使用，以协同糖化褐藻胶。
