Gene Expression and Biochemical Characterization of a GH77 4-α-Glucanotransferase CcGtase From Corallococcus sp. EGB
Gene Expression and Biochemical Characterization of a GH77 4-α-Glucanotransferase CcGtase From Corallococcus sp. EGB
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DOI：
10.1002/star.201800254
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发表时间：
2019-09-01
期刊：
影响因子：
2.3
通讯作者：
Cui Zhongli
中科院分区：
文献类型：
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作者：
Li Zhoukun;Zheng Wenwen;Cui Zhongli
4- α -葡聚糖转移酶催化α -1,4-葡聚糖的分子内和分子间转糖基化，在淀粉代谢和淀粉修饰中起重要作用。本研究克隆了Corallococcus sp. EGB的4- α -葡聚糖转移酶（ccGtase）基因，并在大肠杆菌BL21 （DE3）中表达。CcGtase包含糖苷水解酶家族77 （GH77）的所有保守氨基酸残基，构成了α -淀粉酶家族中酶的活性位点环境，而序列比对表明，来自黏菌的4- α -葡聚糖转移酶与GH77葡聚糖转移酶组成一个独立的基团。产物分析表明，CcGtase能够将麦芽糖低聚糖和淀粉转化为低聚合度（DP < 12）的线性麦芽糖低聚糖，而不是环直链淀粉。对淀粉的转移作用仅在葡萄糖存在的情况下观察到，并且还观察到转移反应延迟产生麦芽糖。在反应过程中，没有观察到明显的水解活性。基于CcGtase序列同源性低（9-35%）和独特的特征，CcGtase可视为GH77家族的新成员。
The enzyme 4-alpha-glucanotransferase catalyzes the intramolecular and intermolecular transglycosylation of alpha-1,4-glucan, which plays an important role in starch metabolism and starch modification. In this study, the gene encoding a 4-alpha-glucanotransferase from Corallococcus sp. EGB (ccGtase) is cloned and expressed in Escherichia coli BL21 (DE3). CcGtase contains all the conserved amino acid residues of glycoside hydrolase family 77 (GH77), which form the active site environment of the enzymes in the alpha-amylase family, while sequence alignments show that 4-alpha-glucanotransferases from myxobacteria constitute an independent group along with the GH77 glucanotransferases. Product analysis shows that CcGtase is able to convert malto-oligosaccharides and starch into linear maltooligosaccharides with a low degree of polymerization (DP < 12) instead of cycloamylose. The transfer action toward starch is only observed in the presence of glucose, and delayed production of maltose from the transfer reaction is also observed. During the reaction, no apparent hydrolytic activity is observed. Based on the low sequence identity (9-35%) and unique characteristics, CcGtase can be regarded as a new member of the GH77 family.