A comparative analysis of two cDNA clones of the cellulase gene family from anaerobic fungus

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题名:	A comparative analysis of two cDNA clones of the cellulase gene family from anaerobic fungus
作者:	Tsai, C.-F.;Liu, J.-H.;Cheng, K.-J.
贡献者:	Department of Biotechnology
关键词:	Cellulase clones;Endoglucanase;Expression;Rumimal fungus
日期:	2003
上传时间:	2010-03-15 08:11:42 (UTC+0)
出版者:	Asia University
摘要:	Cellulase family and some other glycosyl hydrolases of anaerobic fungi inhabiting the digestive tract of ruminants are believed to form an enzyme complex called cellulosome. Study of the individual component of cellulosome may shed light on understanding the organization of this complex and its functional mechanism. We have analysed the primary sequences of two cellulase clones, cel5B and cel6A, isolated from the cDNA library of ruminal fungus, Piromyces rhizinflata strain 2301. The deduced amino acid sequences of the catalytic domain of Cel5B, encoded by cel5B, showed homology with the subfamily 4 of the family 5 (subfamily 5(4)) of glycosyl hydrolases, while cel6A encoded Cel6A belonged to family 6 of glycosyl hydrolases. Phylogenetic tree analysis suggested that the genes of subfamily 5(4) glycosyl hydrolases of P. rhizinflata might have been acquired from rumen bacteria. Cel5B and Cel6A were modular enzymes consisting of a catalytic domain and dockerin domain(s), but not a cellulose binding domain. The occurrence of dockerin domains indicated that both enzymes were cellulosome components. The catalytic domain of the Cel5B (Cel5B′) and Cel6A (Cel6A′) recombinant proteins were purified. The optimal activity conditions with carboxymethyl cellulose (CMC) as the substrate were pH 6.0 and 50°C for Cel5B′, and pH 6.0 and 37–45°C for Cel6A′. Both Cel5B′ and Cel6A′ exhibited activity against CMC, barley β-glucan, Lichenan, and oat spelt xylan. Cel5B′ could also hydrolyse p-nitrophenyl-β-d-cellobioside, Avicel and filter paper while Cel6A′ did not show any activity on these substrates. It is apparent that Cel6A′ acted as an endoglucanase and Cel5B′ possessed both endoglucanase and exoglucanase activities. No synergic effect was observed for these recombinant enzymes in vitro on Avicel and CMC.
關聯:	Piromyces rhizinflata. Anaerobe 9:131-140
显示于类别:	[生物科技學系] 期刊論文

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