Abstract
An endoglycanase gene of Paenibacillus cookii SS-24 was cloned and sequenced. This Pgl8A gene had an open reading frame of 1,230 bp that encoded a putative signal sequence (31 amino acids) and mature enzyme (378 amino acids: 41,835 Da). The enzyme was most homologous to a β-1,3-1,4-glucanase of Bacillus circulans WL-12 with 84% identity. The recombinant enzyme hydrolyzed carboxymethyl cellulose, swollen celluloses, chitosan and lichenan but not Avicel, chitin powder or xylan. With chitosan as the substrate, the optimum temperature and hydrolysis products of the recombinant enzyme varied at pH 4.0 and 8.0. This is the first report that characterizes chitosanase activity under different pH conditions.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Adachi W, Sakihama Y, Shimizu S, Sunami T, Fukazawa T, Suzuki M, Yatsunami R, Nakamura S, Takenaka A (2004) Crystal structure of family GH-8 chitosanase with subclass II specificity from Bacillus sp. K17. J Mol Biol 343:785–795
Choi YJ, Kim EJ, Piao Z, Yun YC, Shin YC (2004) Purification and characterization of chitosanase from Bacillus sp. strain KCTC 0377BP and its application for the production of chitosan oligosaccharides. Appl Environ Microbiol 70:4522–4531
Hash JH, King KW (1958) On the nature of the β-glucosidases of Myrothecium verrucaria. J Biol Chem 232:381–393
Hedges A, Wolfe RS (1974) Extracellular enzyme from Myxobacter AL-1 that exhibits both β-1, 4-glucanase and chitosanase activities. J Bacteriol 120:844–853
Imoto T, Yagishita K (1971) Simple activity measurement of lysozyme. Agri Biol Chem 35:1154–1156
Kimoto H, Kusaoke H, Yamamoto I, Fujii Y, Onodera T, Taketo A (2002) Biochemical and genetic properties of Paenibacillus glycosyl hydrolase having chitosanase activity and discoidin domain. J Biol Chem 277:14695–14702
Mitsutomi M, Isono M, Uchiyama A, Nikaidou N, Ikegami T, Watanabe T (1998) Chitosanase activity of the enzyme previously reported as β-1, 3-1, 4-glucanase from Bacillus circulans WL-12. Biosci Biotechnol Biochem 62:2107–2114
Muzzarelli RAA (1977) Chitin. Pergamon Press, London, p 105
Ogura J, Toyoda A, Kurosawa T, Chong AL, Chohnan S, Masaki T (2006) Purification, characterization, and gene analysis of cellulase (Cel8A) from Lysobacter sp. IB-9374. Biosci Biotechnol Biochem 70:2420–2428
Sakamoto R, Arai M, Murao S (1984) Enzymatic properties of hydrocellulase from Aspergillus aculeatus. J Ferment Technol 62:561–567
Shahidi F, Arachchi JKV, Jeon YJ (1999) Food applications of chitin and chitosans. Trends Food Sci Technol 10:37–51
Shimosaka M, Nogawa M, Ohno Y, Okazaki M (1993) Chitosanase from the plant pathogenic fungus, Fusarium solani f. sp. phaseoli—purification and some properties. Biosci Biotechnol Biochem 57:231–235
Yabuki M, Uchiyama A, Suzuki K, Ando A, Fujii T (1988) Purification and properties of chitosanase from Bacillus circulans MH-K1. J Gen Appl Microbiol 34:255–270
Yatsunami R, Sakihama Y, Suzuki M, Fukazawa T, Shimizu S, Sunami T, Endo K, Takenaka A, Nakamura S (2002) A novel chitosanase from Bacillus sp. strain K17: gene cloning and expression in Escherichia coli. Nucl Acids Res Suppl 2:227–228
Author information
Authors and Affiliations
Corresponding author
Electronic supplementary material
Below is the link to the electronic supplementary material.
10529_2011_759_MOESM2_ESM.docx
Supplementary Fig. 1. SDS–PAGE analysis of the purified enzyme from P. cookii SS-24 and recombinant enzyme. Lane M, protein standard marker (Protein Ladder 10-250 kDa, New England Biolabs); Lane 1, purified enzyme from P. cookii; Lane 2, purified recombinant enzyme. The purified enzymes were applied to 12% SDS–PAGE. (DOCX 94 kb)
10529_2011_759_MOESM3_ESM.docx
Supplementary Fig. 2. Comparison of the amino acid sequence of Pgl8A and other glycosyl hydrolases. The amino acid sequences of GHF-8 enzymes from P. cookii SS-24 (Pgl8A), B. circulans WL-12 (B.WL-12), B. circulans KSM-N257 (B.N257), Lysobacter sp. IB-9374 (L.IB9374), Bacillus sp. KCTC0377BP (B. KCTC) and Bacillus sp. K17 (B.K17) are shown. The conserved amino acid sequence of GHF-8 catalytic module is underlined. Asterisks denote potential catalytic residues. (DOCX 1,305 kb)
Rights and permissions
About this article
Cite this article
Shinoda, S., Kanamasa, S. & Arai, M. Cloning of an endoglycanase gene from Paenibacillus cookii and characterization of the recombinant enzyme. Biotechnol Lett 34, 281–286 (2012). https://doi.org/10.1007/s10529-011-0759-5
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10529-011-0759-5