Cloning, Sequencing and Expression Analysis of the First Cellulase Gene Encoding Cellobiohydrolase 1 from a Cold-adaptive Penicillium chrysogenum FS010
Yunhua HOU
State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China
Department of Food and Biologic Engineering, Shandong Institute of Light Industry, Jinan 250323, China
Search for more papers by this authorCorresponding Author
Tianhong WANG
State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China
*Corresponding author: 86-0531-88566118; Fax, 86-531-88565610; E-mail, [email protected]Search for more papers by this authorHao LONG
State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China
Search for more papers by this authorHuiyuan ZHU
State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China
Department of Food and Biologic Engineering, Shandong Institute of Light Industry, Jinan 250323, China
Search for more papers by this authorYunhua HOU
State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China
Department of Food and Biologic Engineering, Shandong Institute of Light Industry, Jinan 250323, China
Search for more papers by this authorCorresponding Author
Tianhong WANG
State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China
*Corresponding author: 86-0531-88566118; Fax, 86-531-88565610; E-mail, [email protected]Search for more papers by this authorHao LONG
State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China
Search for more papers by this authorHuiyuan ZHU
State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China
Department of Food and Biologic Engineering, Shandong Institute of Light Industry, Jinan 250323, China
Search for more papers by this authorThis work was supported by the grants from the Major State Basic Research Development Program (973) of China (N0. 2003CB716006 and 2004CB719702), the Natural Science Foundation of Shandong Province (No. L2003D01) and the Open Foundation of State Key laboratory of Microbial Technology, Shandong University
Abstract
A cellobiohydrolase 1 gene (cbh1) was cloned from Penicillium chrysogenum FS010 by a modified thermal asymmetric interlaced polymerase chain reaction (TAIL-PCR). DNA sequencing shows that cbh1 has an open reading frame of 1590 bp, encoding a putative protein of 529 amino acid residues. The deduced amino acid sequence revealed that CBHI has a modular structure with a predicted molecular mass of 56 kDa and consists of a fungal type carbohydrate binding module separated from a catalytic domain by a threonine rich linker region. The putative gene product is homologous to fungal cellobiohydrolases in Family 7 of the glycosyl hydrolases. A novel cbh1 promoter (1.3 kb) was also cloned and sequenced, which contains seven putative binding sites (5′-SYGGRG-3′) for the carbon catabolite repressor CRE1. Effect of various carbon sources to the cbh1 transcription of P. chrysogenum was examined by Northern analysis, suggesting that the expression of cbh1 is regulated at transcriptional level. The cbh1 gene in cold-adaptive fungus P. chysogenum was expressed as an active enzyme in Saccharomyces cerevisiae H158. The recombinant CBHI accumulated intracellularly and could not be secreted into the medium.
Edited by Minghua XU
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