从里氏木霉中表达的新型几丁质酶基因的纯化和特性。
Purification and characterization of a novel chitinase gene from Paecilomyces thermophila expressed in Escherichia coli.
机构信息
Department of Biotechnology, College of Food Science and Nutritional Engineering, PO Box 294, China Agricultural University, No. 17 Qinghua Donglu, Haidian District, Beijing 100083, China.
出版信息
Carbohydr Res. 2012 Jan 10;347(1):155-60. doi: 10.1016/j.carres.2011.11.002. Epub 2011 Nov 12.
A novel chitinase gene (PtChiA) from the thermophilic fungus Paecilomyces thermophila was cloned and expressed in Escherichia coli as an intracellular soluble protein. The gene sequence alignment indicates that PtChiA belongs to glycoside hydrolase (GH) family 18 and has an open reading frame comprising of 1473 bp nucleotide sequences with five introns. PtChiA encodes 400 amino acids without any predicted signal peptide. PtChiA was purified by Ni-IDA chromatography. It displayed an acidic optimum pH of 4.5 and broad pH stability (pH 4.0-10.5). The enzyme exhibited an optimal temperature of 50°C and was stable up to 40°C. PtChiA was strongly inhibited by anionic detergent SDS, and also by metal ions Hg(2+) and Mn(2+). It did not exhibit any antifungal activity against pathogenic fungi. It has the ability to hydrolyze colloidal chitin into chito-oligomers suggesting its use in conversion of chitin waste into chito-oligosaccharides.
从嗜热真菌Thermomyces thermophilus中克隆并在大肠杆菌中表达了一种新型几丁质酶基因（PtChiA），作为一种细胞内可溶性蛋白。基因序列比对表明，PtChiA 属于糖苷水解酶（GH）家族 18，具有包含 1473 个核苷酸序列的开放阅读框，其中包含 5 个内含子。PtChiA 编码 400 个氨基酸，没有预测的信号肽。PtChiA 通过 Ni-IDA 层析进行纯化。它显示出酸性最适 pH 值为 4.5，并且具有广泛的 pH 稳定性（pH 4.0-10.5）。该酶的最适温度为 50°C，在 40°C 下稳定。PtChiA 被阴离子洗涤剂 SDS 强烈抑制，也被金属离子 Hg（2+）和 Mn（2+）抑制。它对致病性真菌没有任何抗真菌活性。它具有将胶体几丁质水解成壳寡糖的能力，这表明它可用于将几丁质废物转化为壳寡糖。
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