Cloning of an endoglycanase gene from Paenibacillus cookii and characterization of the recombinant enzyme

Shinoda, Suguru; Kanamasa, Shin; Arai, Motoo

doi:10.1007/s10529-011-0759-5

Cited by 18 publications

(19 citation statements)

References 14 publications

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“…3). The molecular mass of the BGlc8H was similar to those of other GH8 b-glucanases reported (Bueno et al 1990b;Hakamada et al 2002;Ogura et al 2006;Shinoda et al 2012). The molecular masses of GH family 16 (GH16) Bacillus b-1,3-1,4-glucanases (lichenases) were reported to be 25-30 kDa (Planas 2000).…”

Section: Purification Of Bglc8hsupporting

confidence: 70%

“…It showed the highest identity of 98 % to that of the mixed-linked endo-b-1,3-1,4-glucanase (lichenase) of B. circulans WL-12 (P19254), (Bueno et al 1990b), followed by 85 % identity to the cellulase (Pgl8A) of Paenibacillus cookii (AB644221 = BAL46897), (Shinoda et al 2012). Unexpectedly, it showed 81 % identity to the GH8 family chitosanase (Csn1794) of Paenibacillus sp.…”

Section: Resultsmentioning

confidence: 99%

“…1). The highly conserved residues necessary for the catalytic activities of GH8 enzymes have been reported to be one glutamate and one aspartate (Shinoda et al 2012), and the inferred active sites of the endo-b-1,3-1,4-glucanase of B. circulans WL-12 were Glu95 and Asp156 (P19254). The Asp156 was located in the conserved region of ATDGDMDIAYSLLLADKQW.…”

Section: Resultsmentioning

confidence: 99%

“…The closest enzyme was endo-b-1,3-1,4-glucanase (BGC) of B. circulans WL-12 (P19254) that hydrolyzed a mixed-linked b-1,3-1,4-glucan such as lichenan but not CMC (Bueno et al 1990a). Endoglycanase Pgl8A (cellulase) of P. cookii (AB644221 = BAL46897) (Shinoda et al 2012) and chitosanase (Csn1794) of Paenibacillus sp. 1794 (AFP58892) (Zitouni et al 2013) were located near the BGC.…”

Section: Resultsmentioning

confidence: 99%

“…showed much lower CMCase activity (3 %) compared to its chitosanase activity (Choi et al 2004). The GH8 family endoglycanase Pgl8A from P. cookii showed the highest activity toward chitosan, and 34.2 % CMCase activity and 9 % lichenase activity compared to the chitosanase activity (data for other b-glucans were not available) (Shinoda et al 2012). The GH8 family chitosanase Csn1794 from Paenibacillus sp.…”

Section: Substrate Specificity Of Bglc8hmentioning

confidence: 99%

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Characterization of a GH family 8 β-1,3-1,4-glucanase with distinctive broad substrate specificity from Paenibacillus sp. X4

Jung

Jeong

et al. 2014

Biotechnol Lett

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A β-1,3-1,4 glucanase gene of Paenibacillus sp. X4, bglc8H, was cloned and characterized. BGlc8H was predicted to be a protein of 409 amino acid residues, including a signal peptide of 31 amino acids. The mature enzyme was predicted to have 378 amino acid residues; its [corrected] molecular mass and pI were estimated as 41,561 Da and 7.61, respectively. BGlc8H belongs to glycoside hydrolase family 8 (GH8). Site-directed mutants of Glu95 and Asp156 of BGlc8H showed a near-complete loss of activity, indicating that they are catalytically-active residues. Unlike other GH8 members, BGlc8H had broad substrate specificity and hydrolyzed barley-β-D-glucan > chitosan > carboxymethyl-cellulose > and lichenan. BGlc8H had a lower ratio of lichenase/barley-β-D-glucanase activities compared to GH16 enzymes. BGlc8H was optimally active at pH 5 and 50 °C, except for barley-β-D-glucanase (40 °C) and chitosanase (pH 7) activities. BGlc8H hydrolyzed cello-oligosaccharides (G3-G6) to G3 and G2 but not to G1. Ca(2+) increased the activity and thermostability of BGlc8H for lichenan suggesting its use for the saccharification of cellulosic biomass.

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Section: Purification Of Bglc8hsupporting

confidence: 70%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Substrate Specificity Of Bglc8hmentioning

confidence: 99%

See 3 more Smart Citations

Characterization of a GH family 8 β-1,3-1,4-glucanase with distinctive broad substrate specificity from Paenibacillus sp. X4

Jung

Jeong

et al. 2014

Biotechnol Lett

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New Paradigm in Degradation of Lignocellulosic Biomass and Discovery of Novel Microbial Strains

Rai

Agrawal

Chadha

2019

Microbial Diversity in Ecosystem Sustainability and Biotechnological Applications

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Biochemical and molecular characterization of a thermostable chitosanase produced by the strain Paenibacillus sp. 1794 newly isolated from compost

Zitouni

Fortin

Scheerle

et al. 2012

Appl Microbiol Biotechnol

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Chitosan raises a great interest among biotechnologists due to its potential for applications in biomedical or environmental fields. Enzymatic hydrolysis of chitosan is a recognized method allowing control of its molecular size, making possible its optimization for a given application. During the industrial hydrolysis process of chitosan, viscosity is a major problem; which can be circumvented by raising the temperature of the chitosan solution. A thermostable chitosanase is compatible with enzymatic hydrolysis at higher temperatures thus allowing chitosan to be dissolved at higher concentrations. Following an extensive micro-plate screening of microbial isolates from various batches of shrimp shells compost, the strain 1794 was characterized and shown to produce a thermostable chitosanase. The isolate was identified as a novel member of the genus Paenibacillus, based on partial 16S rDNA and rpoB gene sequences. Using the chitosanase (Csn1794) produced by this strain, a linear time course of chitosan hydrolysis has been observed for at least 6 h at 70 °C. Csn1794 was purified and its molecular weight was estimated at 40 kDa by SDS-PAGE. Optimum pH was about 4.8, the apparent Km and the catalytic constant kcat were 0.042 mg/ml and 7,588 min⁻¹, respectively. The half-life of Csn1794 at 70 °C in the presence of chitosan substrate was >20 h. The activity of chitosanase 1794 varied little with the degree of N-acetylation of chitosan. The enzyme also hydrolyzed carboxymethylcellulose but not chitin. Chitosan or cellulose-derived hexasaccharides were cleaved preferentially in a symmetrical way ("3+3") but hydrolysis rate was much faster for (GlcN)₆ than (Glc)₆. Gene cloning and sequencing revealed that Csn1794 belongs to family 8 of glycoside hydrolases. The enzyme should be useful in biotechnological applications of chitosan hydrolysis, dealing with concentrated chitosan solutions at high temperatures.

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Cloning of an endoglycanase gene from Paenibacillus cookii and characterization of the recombinant enzyme

Cited by 18 publications

References 14 publications

Characterization of a GH family 8 β-1,3-1,4-glucanase with distinctive broad substrate specificity from Paenibacillus sp. X4

Characterization of a GH family 8 β-1,3-1,4-glucanase with distinctive broad substrate specificity from Paenibacillus sp. X4

New Paradigm in Degradation of Lignocellulosic Biomass and Discovery of Novel Microbial Strains

Biochemical and molecular characterization of a thermostable chitosanase produced by the strain Paenibacillus sp. 1794 newly isolated from compost

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