A novel carboxyl-terminal protease derived from Paenibacillus lautusCHN26 exhibiting high activities at multiple sites of substrates

BMC Biotechnology

Research Article

Qunxin She¹  Yunxia Li²  Yingjie Pan²  Lanming Chen² 

[1] Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, DK-2200N, Copenhagen, Denmark;Key Laboratory of Quality and Safety Risk Assessment for Aquatic Products on Storage and Preservation (Shanghai), China Ministry of Agriculture, Engineering Centre for Quality Control and Risk Assessment of Aquatic Products, College of Food Science and Technology, Shanghai Ocean University, 999 Hu Cheng Huan Road, 201306, Shanghai, People's Republic of China;

关键词: CtpA;    Paenibacillus;    Protease;    Aqueous environment;   

DOI  :  10.1186/1472-6750-13-89

 received in 2013-08-12, accepted in 2013-10-17,  发布年份 2013

来源: Springer

【 摘 要 】

BackgroundCarboxyl-terminal protease (CtpA) plays essential functions in posttranslational protein processing in prokaryotic and eukaryotic cells. To date, only a few bacterial ctpA genes have been characterized. Here we cloned and characterized a novel CtpA. The encoding gene, ctpAp (ctpA of Paenibacillus lautus), was derived from P. lautus CHN26, a Gram-positive bacterium isolated by functional screening. Recombinant protein was obtained from protein over-expression in Escherichia coli and the biochemical properties of the enzyme were investigated.ResultsScreening of environmental sediment samples with a skim milk-containing medium led to the isolation of a P. lautus CHN26 strain that exhibited a high proteolytic activity. A gene encoding a carboxyl-terminal protease (ctpAp) was cloned from the isolate and characterized. The deduced mature protein contains 466 aa with a calculated molecular mass of 51.94 kDa, displaying 29-38% amino acid sequence identity to characterized bacterial CtpA enzymes. CtpAp contains an unusual catalytic dyad (Ser309-Lys334) and a PDZ substrate-binding motif, characteristic for carboxyl-terminal proteases. CtpAp was expressed as a recombinant protein and characterized. The purified enzyme showed an endopeptidase activity, which effectively cleaved α S1- and β- casein substrates at carboxyl-terminus as well as at multiple internal sites. Furthermore, CtpAp exhibited a high activity at room temperature and strong tolerance to conventional protease inhibitors, demonstrating that CtpAp is a novel endopeptidase.ConclusionsOur work on CtpA represents the first investigation of a member of Family II CtpA enzymes. The gene was derived from a newly isolated P. lautus CHN26 strain exhibiting a high protease activity in the skim milk assay. We have demonstrated that CtpAp is a novel endopeptidase with distinct cleavage specificities, showing a strong potential in biotechnology and industry applications.

【 授权许可】

Unknown   
© Li et al.; licensee BioMed Central Ltd. 2013. This article is published under license to BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.