Molecular cloning, sequencing and expression of the gene encoding a novel chitinase A from a marine bacterium, Pseudomonas sp PE2, and its domain structure
Molecular cloning, sequencing and expression of the gene encoding a novel chitinase A from a marine bacterium, Pseudomonas sp PE2, and its domain structure
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DOI：
10.1007/s00253-002-1154-x
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发表时间：
2003-04-01
影响因子：
5
通讯作者：
Kamei, Y
中科院分区：
文献类型：
--
作者：
Kitamura, E;Kamei, Y
从一株能降解卟啉腐霉细胞壁的海洋细菌Pseudomonassp.PE2中克隆了编码几丁质酶A（PchA）的基因。推测的PchA是一个由N端信号肽、糖苷水解酶家族18催化结构域、几丁质结合结构域（ChBD）和碳水化合物结合模块（CBM）组成的模块化酶。ChBD（PchA）的氨基酸序列在CBM家族12中是高度保守的，CBM家族12也容纳没有AKWWTQG基序的ChBD，AKWWTQG基序是细菌几丁质酶和灰色链霉菌蛋白酶C中常见的结构域。有趣的是，CBMPchA显示出与来自混合纤维弧菌（Cellvibrio mixtus）的内切葡聚糖酶B的C-末端区域的显著序列同源性，所述内切葡聚糖酶是CBM家族6的成员。这是首次报道的几丁质酶具有与CBM家族6高度相似的结构域。缺失分析表明，ChBDPchA可能在天然甲壳素和壳聚糖的结合中发挥重要作用，而不是加工甲壳素。CBMPchA也似乎在木聚糖和Avicel的结合中发挥这样的作用。这些结果表明，PchA的C-末端区域可能是一个关键的组成部分，在结合几丁质的细胞壁的紫菜或其他海洋生物的结构组成部分。
The pchA gene encoding chitinase A (PchA) from a Pythium porphyrae cell-wall-degrading marine bacterium, Pseudomonas sp. PE2, was cloned and characterized. The deduced PchA was a modular enzyme composed of an N-terminal signal peptide, a glycoside hydrolase family 18 catalytic domain that was responsible for the chitinase activity, the chitin-binding domains (ChBDs), and the carbohydrate-binding modules (CBM). The amino acid sequence of ChBD(PchA) was highly conserved in the CBM family 12 that also accommodates ChBDs without an AKWWTQG motif, a domain commonly found in bacterial chitinase and Streptomyces griseus protease C. Interestingly, CBMPchA showed significant sequence homology to the C-terminal region of endoglucanase B from Cellvibrio mixtus, which is a member of CBM family 6. This is the first report of a chitinase possessing a domain with high similarity to CBM family 6. Deletion analysis indicated clearly that ChBDPchA might play an important role in the binding of native chitin and chitosan, but not processed chitin. CBMPchA also appeared to play such a role in the binding of xylan and Avicel. These results suggest that the C-terminal region of PchA might be a key component in the binding of chitin in the cell walls of P. porphyrae or other structural components of marine organisms.