Cloning, DNA sequencing, and expression of the gene encoding Clostridium thermocellum cellulase CelJ, the largest catalytic component of the cellulosome
Cloning, DNA sequencing, and expression of the gene encoding Clostridium thermocellum cellulase CelJ, the largest catalytic component of the cellulosome
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DOI：
10.1128/jb.178.19.5732-5740.1996
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发表时间：
1996-10-01
影响因子：
3.2
通讯作者：
Ohmiya, K
中科院分区：
文献类型：
--
作者：
Ahsan, MM;Kimura, T;Ohmiya, K
热纤梭菌F1 celJ基因编码内切葡聚糖酶J（CelJ），由4,803个核苷酸的开放阅读框（ORF）组成，编码1,601个氨基酸、分子量为178,055的蛋白质。通过与截短的 CelJ 衍生物的 N 端序列比较，确认 ORF 是细胞。 CelJ是一种模块化酶，由N端信号肽和按以下顺序排列的六个结构域组成：S层同源结构域、未知功能结构域(UD-1)、亚家族E1内切葡聚糖酶结构域、家族J内切葡聚糖酶结构域、对接结构域和另一个未知功能结构域(UD-2)。 UD-1与UD-2没有显着相似性。 CelJ 水解羧甲基纤维素和木聚糖，木聚糖酶活性归因于家族 J 结构域。针对截短的 CelJ 产生的抗血清与热纤梭菌 F1 的多纤维素酶体中所含的蛋白质发生交叉反应。这些结果强烈表明，CelJ 相当于 S-2，S-2 被确定为热纤梭菌 YS 的纤维素体中最大的催化成分。第二个但不完整的 ORF 编码属于 E2 内切葡聚糖酶亚家族的酶，位于 celJ 下游。
The Clostridium thermocellum F1 celJ gene, encoding endoglucanase J (CelJ), consists of an open reading frame (ORF) of 4,803 nucleotides and encodes a protein of 1,601 amino acids with a molecular weight of 178,055. The ORF was confirmed as cell by comparison with the N-terminal sequence of a truncated CelJ derivative. CelJ is a modular enzyme composed of N-terminal signal peptide and six domains in the following order: an S-layer homology domain, a domain of unknown function (UD-1), a subfamily El endoglucanase domain, a family J endoglucanase domain, a docking domain, and another domain of unknown function (UD-2). UD-1 has no significant similarity to UD-2. CelJ hydrolyzed carboxymethylcellulose and xylan, and xylanase activity was ascribed to the family J domain. Antiserum raised against the truncated CelJ crossreacted with proteins contained in the cellulosome of C. thermocellum F1. These results strongly suggest that CelJ is equivalent to S-2, which was identified as the largest catalytic component in the cellulosome of C. thermocellum YS. A second but incomplete ORF encoding an enzyme classified in subfamily E2 endoglucanase, was located downstream of celJ.