文献类型：期刊文献

英文题名：Development of a novel Aspergillus uracil deficient expression system and its application in expressing a cold-adapted α-amylase gene from Antarctic fungi Geomyces pannorum

作者：Mao, Youzhi[1];Yin, Yanchen[1];Zhang, Lujia[1];Alias, Siti Aisyah[2];Gao, Bei[1];Wei, Dongzhi[1]

机构：[1]E China Univ Sci & Technol, New World Inst Biotechnol, State Key Lab Bioreactor Engn, Shanghai 200237, Peoples R China;[2]Univ Malaya, Inst Ocean & Earth Sci, Inst Postgrad Studies, Kuala Lumpur 50603, Malaysia

年份：2015

卷号：50

期号：10

起止页码：1581

外文期刊名：PROCESS BIOCHEMISTRY

收录：;EI(收录号：20152801016292);WOS:【SCI-EXPANDED(收录号:WOS:000361775400011)】；

基金：This work was funded by The National Natural Foundation of China (No. 31201296), the Fundamental Research Funds for the Central Universities, and the National High Technology Research and Development Program of China (No.2013AA102109). We are grateful to Institute of Ocean and Earth Sciences, University of Malaya for offering strain Geomyces pannorum. This polar work was supported by Higher Institute Centre of Excellence Grant, Ministry Of Education (No. IOES-2014G).

语种：英文

外文关键词：Cold-adapted alpha-amylase; pyrF; Aspergillus oryzae; Geomyces pannorum; Heterologous expression

摘要：Filamentous fungi have increasingly been used as hosts for heterologous protein production because of their high secretion capability and ability to add eukaryotic post-translational modifications. In this study, a novel uracil-deficient Aspergillus transformation system, which was based on an orotate phosphoribosyltransferase (pyrF) nutritional selection marker, was discovered. Additionally, a universal, purify-able vector that directed genes into the Aspergillus host strain was engineered. A genomic DNA segment encoding a novel a-amylase was isolated from the psychrotolerant fungus Geomyces pannorum and the open reading frame was determined, deduced 497 amino acids. G. pannorum alpha-amylase was then expressed in the newly constructed Aspergillus oryzae system, with an amylase activity reaching 958 U/ml. It was purified to electrophoretic homogeneity and has a molecular mass of approximately 54 kDa. The enzyme exhibited an optimal activity at pH 5.0 and 40 degrees C and retained over 20% of maximal activity over the temperature range 0-20 degrees C. To our knowledge, this report is the first of the heterologous expression of a cold-adapted enzyme in filamentous fungi. G. pannorum alpha-amylase is an economical amylase with many potential applications. (C) 2015 Elsevier Ltd. All rights reserved.