Comparative Analyses of Two Thermophilic Enzymes Exhibiting both β-1,4 Mannosidic and β-1,4 Glucosidic Cleavage Activities from Caldanaerobius polysaccharolyticus
Comparative Analyses of Two Thermophilic Enzymes Exhibiting both β-1,4 Mannosidic and β-1,4 Glucosidic Cleavage Activities from Caldanaerobius polysaccharolyticus
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DOI：
10.1128/jb.00257-10
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发表时间：
2010-08-01
影响因子：
3.2
通讯作者：
Cann, Isaac K. O.
中科院分区：
文献类型：
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作者：
Han, Yejun;Dodd, Dylan;Cann, Isaac K. O.
含有甘露聚糖的多糖的水解需要内切-1,4- β -甘露聚糖酶和1,4- β -甘露聚糖苷酶的活性。本文研究了水解Caldanaerobius多糖的两种内切-1,4-甘露聚糖酶（Man5A和Man5B）的生化特性。Man5A由一个n端信号肽（SP）、一个催化结构域、两个碳水化合物结合模块（CBMs）和三个表面层同源重复序列组成，而Man5B缺乏SP、CBMs和SLH重复序列。为了深入了解这两种糖苷水解酶家族5 （GH5）酶如何帮助细菌获取能量，以及这两种酶在生物燃料工业中的潜在应用，我们对Man5A的两种衍生物（Man5A-TM1 [TM1代表截断突变体1]，缺乏SP和SLH重复序列，以及Man5A- tm2，缺乏SP、CBMs和SLH重复序列）和野生型Man5B进行了生化分析。Man5A衍生物具有内切-1,4- β甘露聚糖酶和内切-1,4- β -葡聚糖酶活性，水解的寡糖聚合度（DP）为4或更高。Man5B具有内切-1,4-甘露聚糖酶活性，内切-1,4- β -葡聚糖酶活性较弱；然而，该酶也表现出1,4-甘露糖苷酶和纤维素糊精酶的活性。Man5A- tm1对可溶性和不溶性多糖的催化活性均高于Man5A- tm2和Man5B，表明CBMs增强了Man5A的催化活性。此外，Man5A-TM1与Man5B协同水解-甘露聚糖和羧甲基纤维素。因此，这两种酶的多功能性使它们成为生物燃料工业中含甘露聚糖多糖解聚的资源。此外，根据生物化学和基因组学数据，提出了溶多糖酵母利用含甘露聚糖营养物质的分子机制。
The hydrolysis of polysaccharides containing mannan requires endo-1,4-beta-mannanase and 1,4-beta-mannosidase activities. In the current report, the biochemical properties of two endo-beta-1,4-mannanases (Man5A and Man5B) from Caldanaerobius polysaccharolyticus were studied. Man5A is composed of an N-terminal signal peptide (SP), a catalytic domain, two carbohydrate-binding modules (CBMs), and three surface layer homology (SLH) repeats, whereas Man5B lacks the SP, CBMs, and SLH repeats. To gain insights into how the two glycoside hydrolase family 5 (GH5) enzymes may aid the bacterium in energy acquisition and also the potential application of the two enzymes in the biofuel industry, two derivatives of Man5A (Man5A-TM1 [TM1 stands for truncational mutant 1], which lacks the SP and SLH repeats, and Man5A-TM2, which lacks the SP, CBMs, and SLH repeats) and the wild-type Man5B were biochemically analyzed. The Man5A derivatives displayed endo-1,4-beta-mannanase and endo-1,4-beta-glucanase activities and hydrolyzed oligosaccharides with a degree of polymerization (DP) of 4 or higher. Man5B exhibited endo-1,4-beta-mannanase activity and little endo-1,4-beta-glucanase activity; however, this enzyme also exhibited 1,4-beta-mannosidase and cellodextrinase activities. Man5A-TM1, compared to either Man5A-TM2 or Man5B, had higher catalytic activity with soluble and insoluble polysaccharides, indicating that the CBMs enhance catalysis of Man5A. Furthermore, Man5A-TM1 acted synergistically with Man5B in the hydrolysis of beta-mannan and carboxymethyl cellulose. The versatility of the two enzymes, therefore, makes them a resource for depolymerization of mannan-containing polysaccharides in the biofuel industry. Furthermore, on the basis of the biochemical and genomic data, a molecular mechanism for utilization of mannan-containing nutrients by C. polysaccharolyticus is proposed.