Department of Biotechnology, College of Food Science and Nutritional Engineering, China Agricultural University, PO Box 294, No.17 Qinghua Donglu, Haidian District, Beijing, 100083, China.
Appl Biochem Biotechnol. 2013 May;170(2):420-35. doi: 10.1007/s12010-013-0198-y. Epub 2013 Mar 29.
A novel α-amylase (McAmyA) from the thermophilic fungus, Malbranchea cinnamomea was purified, characterized and crystallized in the present study. McAmyA was purified to apparent homogeneity with a molecular mass of 60.3 kDa on SDS-PAGE. The enzyme exhibited maximal activity at pH 6.5 and was stable within pH 5.0-10.0. It was most active at 65 °C and was stable up to 50 °C. McAmyA was capable of hydrolyzing amylose, starch, amylopectin, pullulan, cyclodextrins and maltooligosaccharides. The full-length cDNA of an α-amylase gene (McAmyA) from the strain was cloned. McAmyA consisted of a 1,476-bp open reading frame encoding 492 amino acids. It displayed the highest amino acid sequence homology (less than 60 %) with the reported α-amylases. The crystal structure of McAmyA was solved at a resolution of 2.25 Å (PDB code 3VM7). The overall structure of McAmyA reveals three domains with ten α helices and 14 β strands, and the putative catalytic residues are positioned at domain A with somewhat different secondary structural circumstances compared with typical α-amylases.
本研究对来自嗜热真菌 Malbranchea cinnamomea 的新型α-淀粉酶（McAmyA）进行了纯化、表征和结晶。McAmyA 在 SDS-PAGE 上显示出约 60.3 kDa 的表观分子量，可达到明显的均一性。该酶在 pH 6.5 时表现出最大活性，在 pH 5.0-10.0 范围内稳定。它在 65°C 时最活跃，在 50°C 下稳定。McAmyA 能够水解直链淀粉、淀粉、支链淀粉、普鲁兰、环糊精和麦芽寡糖。该菌株的全长 α-淀粉酶基因（McAmyA）的 cDNA 被克隆。McAmyA 由一个 1476 个碱基对的开放阅读框编码 492 个氨基酸。它与报道的 α-淀粉酶的氨基酸序列同源性最高（低于 60%）。McAmyA 的晶体结构在 2.25 Å 的分辨率下得到解决（PDB 代码 3VM7）。McAmyA 的整体结构揭示了三个结构域，包含十个α螺旋和 14 个β链，与典型的α-淀粉酶相比，假定的催化残基位于结构域 A 中，其二级结构环境略有不同。
