• Journal of Microbiology
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• v.34 no.4
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• pp.305-310
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• 1996

A microorganism producing carboxymethyl-cellulase (CMCase) was isolated from 300 soil and compost samples. The isolate was identified as Bacillus sp. by $Biolog^{TM}$ test and fatty acid analysis, and named as Bacillus sp. KD1014. The isolate could degrade, in addition to CMC, various kinds of polysaccharides such as levan, xylan, starch, and filter paper but hardly degrade microcrystalline Avicel. The optimum growth and CMCase production of the isolate was observed between 16-and 25 hr-culture at 45$^{\circ}C$ and pH 5.0. The maximum CMCase activity was observed at pH 4.5 and 6$0^{\circ}C$. The CMCase was found to bind to Avicel. The CMCase was internally cleaved as growth continued. When crude supernatant was used for activity staining, three major bands were detected on a native gel, however, only one major band was detected on a denaturating gel after removal of the detergent.

• Journal of Applied Biological Chemistry
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• v.42 no.3
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• pp.107-112
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• 1999

A carboxymethyl-cellulase (CMCase) was purified from the culture supernatant of Bacillus sp. KD1014 by ultrafiltration, ammonium sulfate precipitation, and a series of chromatography on QAE-Sephadex A-50, hydroxylapatite and Sephadex G-75. The purified CMCase was a single protein of 32 kDa, showed an optimum activity at $60^{\circ}C$ and pH 6.0, and had a half-life of 23 min at $70^{\circ}C$. The enzyme activity was not influenced by metal ions such as $Mg^{2+},\;Fe^{3+},\;K^+,\;Zn^{2+}$, and $Cu^{2+}$ at a concentration of 1.0 mM, partially inhibited by $Mn^{2+}$ and $Ag^+$, and significantly inhibited by pentachlorophenol (PCP). The purified enzyme showed a 3.9-times higher activity on lichenan than on CMC, but hardly cleaved xylan, starch, avicel, laminarin, filter paper and levan. The results of activity staining of the purified enzyme separated by native and denaturing gel electrophoresis suggested that the CMCase might exist in dimeric, oligomeric or aggregated form as well as in monomeric form. The enzymatic cleavage products from cellotetraose indicated that the CMCase possessed transglycosylation activity.

• Journal of Microbiology and Biotechnology
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• v.28 no.12
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• pp.2036-2045
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• 2018

An endo-${\beta}$-1,4-glucanase gene, cel5L, was cloned using the shot-gun method from Bacillus sp.. The gene, which contained a predicted signal peptide, encoded a protein of 496 amino acid residues, and the molecular mass of the mature Cel5L was estimated to be 51.8 kDa. Cel5L contained a catalytic domain of glycoside hydrolase (GH) family 5 and a carbohydrate-binding module family 3 (CBM_3). Chromatography using HiTrap Q and CHT-II resulted in the isolation of two truncated forms corresponding to 50 (Cel5L-p50) and 35 kDa (Cel5L-p35, CBM_3-deleted form). Both enzymes were optimally active at pH 4.5 and $55^{\circ}C$, but had different half-lives of 4.0 and 22.8 min, respectively, at $70^{\circ}C$. The relative activities of Cel5L-p50 and Cel5L-p35 for barley ${\beta}$-glucan were 377.0 and 246.7%, respectively, compared to those for carboxymethyl-cellulose. The affinity and hydrolysis rate of pNPC by Cel5L-p35 were 1.7 and 3.3 times higher, respectively, than those by Cel5L-p50. Additions of each to a commercial enzyme set increased saccharification of pretreated rice straw powder by 17.5 and 21.0%, respectively. These results suggest CBM_3 is significantly contributing to thermostability, and to affinity and substrate specificity for small substrates, and that these two enzymes could be used as additives to enhance enzymatic saccharification.