来自普拉宁毛霉的新型葡萄糖脱氢酶：纯化、特性鉴定、分子克隆及在酱油曲霉中的基因表达

Novel glucose dehydrogenase from Mucor prainii: Purification, characterization, molecular cloning and gene expression in Aspergillus sojae.

作者信息

Satake Ryoko, Ichiyanagi Atsushi, Ichikawa Keiichi, Hirokawa Kozo, Araki Yasuko, Yoshimura Taro, Gomi Keiko

机构信息

Research and Development Division, Kikkoman Corporation, 399 Noda, Chiba 278-0037, Japan.

出版信息

J Biosci Bioeng. 2015 Nov;120(5):498-503. doi: 10.1016/j.jbiosc.2015.03.012. Epub 2015 Apr 23.

DOI:10.1016/j.jbiosc.2015.03.012

PMID:25912449

Abstract

Glucose dehydrogenase (GDH) is of interest for its potential applications in the field of glucose sensors. To improve the performance of glucose sensors, GDH is required to have strict substrate specificity. A novel flavin adenine dinucleotide (FAD)-dependent GDH was isolated from Mucor prainii NISL0103 and its enzymatic properties were characterized. This FAD-dependent GDH (MpGDH) exhibited high specificity toward glucose. High specificity for glucose was also observed even in the presence of saccharides such as maltose, galactose and xylose. The molecular masses of the glycoforms of GDH ranged from 90 to 130 kDa. After deglycosylation, a single 80 kDa band was observed. The gene encoding MpGDH was cloned and expressed in Aspergillus sojae. The apparent kcat and Km values of recombinant enzyme for glucose were found to be 749.7 s(-1) and 28.3 mM, respectively. The results indicated that the characteristics of MpGDH were suitable for assaying blood glucose levels.

摘要

葡萄糖脱氢酶（GDH）因其在葡萄糖传感器领域的潜在应用而备受关注。为了提高葡萄糖传感器的性能，要求GDH具有严格的底物特异性。从普拉宁毛霉NISL0103中分离出一种新型的黄素腺嘌呤二核苷酸（FAD）依赖性GDH，并对其酶学性质进行了表征。这种FAD依赖性GDH（MpGDH）对葡萄糖表现出高特异性。即使在存在麦芽糖、半乳糖和木糖等糖类的情况下，也观察到对葡萄糖的高特异性。GDH糖型的分子量范围为90至130 kDa。去糖基化后，观察到一条单一的80 kDa条带。克隆了编码MpGDH的基因并在大豆曲霉中表达。发现重组酶对葡萄糖的表观kcat和Km值分别为749.7 s(-1)和28.3 mM。结果表明，MpGDH的特性适用于测定血糖水平。