Effect of Family 22 Carbohydrate-Binding Module on the Thermostability of Xyn10B Catalytic Module fromClostridium stercorarium

2006 ◽  
Vol 70 (12) ◽  
pp. 3039-3041 ◽  
Author(s):  
Rie ARAKI ◽  
Shuichi KARITA ◽  
Akiyoshi TANAKA ◽  
Tetsuya KIMURA ◽  
Kazuo SAKKA
Keyword(s):  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
Catalytic Module

scholarly journals Modular Glucuronoxylan-Specific Xylanase with a Family CBM35 Carbohydrate-Binding Module

2012 ◽  
Vol 78 (11) ◽  
pp. 3923-3931 ◽  
Author(s):  
Susana Valeria Valenzuela ◽  
Pilar Diaz ◽  
F. I. Javier Pastor
Keyword(s):  
Catalytic Activity ◽  
Kinetic Parameters ◽  
Glucuronic Acid ◽  
Specific Activity ◽  
Dimensional Structure ◽  
Titration Calorimetry ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
Content Type ◽  
Catalytic Module

ABSTRACTXyn30D from the xylanolytic strainPaenibacillus barcinonensishas been identified and characterized. The enzyme shows a modular structure comprising a catalytic module family 30 (GH30) and a carbohydrate-binding module family 35 (CBM35). Like GH30 xylanases, recombinant Xyn30D efficiently hydrolyzed glucuronoxylans and methyl-glucuronic acid branched xylooligosaccharides but showed no catalytic activity on arabinose-substituted xylans. Kinetic parameters of Xyn30D were determined on beechwood xylan, showing aKmof 14.72 mg/ml and akcatvalue of 1,510 min−1. The multidomain structure of Xyn30D clearly distinguishes it from the GH30 xylanases characterized to date, which are single-domain enzymes. The modules of the enzyme were individually expressed in a recombinant host and characterized. The isolated GH30 catalytic module showed specific activity, mode of action on xylan, and kinetic parameters that were similar to those of the full-length enzyme. Computer modeling of the three-dimensional structure of Xyn30D showed that the catalytic module is comprised of a common (β/α)8barrel linked to a side-associated β-structure. Several derivatives of the catalytic module with decreasing deletions of this associated structure were constructed. None of them showed catalytic activity, indicating the importance of the side β-structure in the catalysis of Xyn30D. Binding properties of the isolated carbohydrate-binding module were analyzed by affinity gel electrophoresis, which showed that the CBM35 of the enzyme binds to soluble glucuronoxylans and arabinoxylans. Analysis by isothermal titration calorimetry showed that CBM35 binds to glucuronic acid and requires calcium ions for binding. Occurrence of a CBM35 in a glucuronoxylan-specific xylanase is a differential trait of the enzyme characterized.

scholarly journals Domain Analysis of a Modular α-l-Arabinofuranosidase with a Unique Carbohydrate Binding Strategy from the Fiber-Degrading Bacterium Fibrobacter succinogenes S85

2010 ◽  
Vol 192 (20) ◽  
pp. 5424-5436 ◽  
Author(s):  
Shosuke Yoshida ◽  
Charles W. Hespen ◽  
Robert L. Beverly ◽  
Roderick I. Mackie ◽  
Isaac K. O. Cann
Keyword(s):  
Plant Cell Wall ◽  
Catalytic Efficiency ◽  
Glycoside Hydrolases ◽  
Site Directed Mutagenesis ◽  
Biofuel Production ◽  
Carbohydrate Binding ◽  
Fibrobacter Succinogenes ◽  
Carbohydrate Binding Module ◽  
Degrading Bacterium ◽  
Catalytic Module

ABSTRACT Family 43 glycoside hydrolases (GH43s) are known to exhibit various activities involved in hemicellulose hydrolysis. Thus, these enzymes contribute to efficient plant cell wall degradation, a topic of much interest for biofuel production. In this study, we characterized a unique GH43 protein from Fibrobacter succinogenes S85. The recombinant protein showed α-l-arabinofuranosidase activity, specifically with arabinoxylan. The enzyme is, therefore, an arabinoxylan arabinofuranohydrolase (AXH). The F. succinogenes AXH (FSUAXH1) is a modular protein that is composed of a signal peptide, a GH43 catalytic module, a unique β-sandwich module (XX domain), a family 6 carbohydrate-binding module (CBM6), and F. succinogenes-specific paralogous module 1 (FPm-1). Truncational analysis and site-directed mutagenesis of the protein revealed that the GH43 domain/XX domain constitute a new form of carbohydrate-binding module and that residue Y484 in the XX domain is essential for binding to arabinoxylan, although protein structural analyses may be required to confirm some of the observations. Kinetic studies demonstrated that the Y484A mutation leads to a higher k cat for a truncated derivative of FSUAXH1 composed of only the GH43 catalytic module and the XX domain. However, an increase in the Km for arabinoxylan led to a 3-fold decrease in catalytic efficiency. Based on the knowledge that most XX domains are found only in GH43 proteins, the evolutionary relationships within the GH43 family were investigated. These analyses showed that in GH43 members with a XX domain, the two modules have coevolved and that the length of a loop within the XX domain may serve as an important determinant of substrate specificity.

scholarly journals A mannanase, ManA, of the polycentric anaerobic fungusOrpinomycessp. strain PC-2 has carbohydrate binding and docking modules

2005 ◽  
Vol 51 (7) ◽  
pp. 559-568 ◽  
Author(s):  
Eduardo A Ximenes ◽  
Huizhong Chen ◽  
Irina A Kataeva ◽  
Michael A Cotta ◽  
Carlos R Felix ◽  
...  
Keyword(s):  
Specific Activity ◽  
High Specific Activity ◽  
Glycoside Hydrolases ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
Amino Acid Residues ◽  
Anaerobic Fungus ◽  
Ph Profile ◽  
Catalytic Module ◽  
Cellulose Binding

The anaerobic fungus Orpinomyces sp. strain PC-2 produces a broad spectrum of glycoside hydrolases, most of which are components of a high molecular mass cellulosomal complex. Here we report about a cDNA (manA) having 1924 bp isolated from the fungus and found to encode a polypeptide of 579 amino acid residues. Analysis of the deduced sequence revealed that it had a mannanase catalytic module, a family 1 carbohydrate-binding module, and a noncatalytic docking module. The catalytic module was homologous to aerobic fungal mannanases belonging to family 5 glycoside hydrolases, but unrelated to the previously isolated mannanases (family 26) of the anaerobic fungus Piromyces. No mannanase activity could be detected in Escherichia coli harboring a manA-containing plasmid. The manA was expressed in Saccharomyces cerevisiae and ManA was secreted into the culture medium in multiple forms. The purified extracellular heterologous mannanase hydrolyzed several types of mannan but lacked activity against cellulose, chitin, or β-glucan. The enzyme had high specific activity toward locust bean mannan and an extremely broad pH profile. It was stable for several hours at 50 °C, but was rapidly inactivated at 60 °C. The carbohydrate-binding module of the Man A produced separately in E. coli bound preferably to insoluble lignocellulosic substrates, suggesting that it might play an important role in the complex enzyme system of the fungus for lignocellulose degradation.Key words: Orpinomyces, anaerobic fungi, mannanase, cellulose-binding module, cellulosome.

scholarly journals Characterization of a Cellulase Containing a Family 30 Carbohydrate-Binding Module (CBM) Derived from Clostridium thermocellum CelJ: Importance of the CBM to Cellulose Hydrolysis

2003 ◽  
Vol 185 (2) ◽  
pp. 504-512 ◽  
Author(s):  
Takamitsu Arai ◽  
Rie Araki ◽  
Akiyoshi Tanaka ◽  
Shuichi Karita ◽  
Tetsuya Kimura ◽  
...  
Keyword(s):  
Clostridium Thermocellum ◽  
Tertiary Structure ◽  
Cellulose Hydrolysis ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
Strong Activity ◽  
Catalytic Function ◽  
Catalytic Module ◽  
Calorimetric Studies

ABSTRACT Clostridium thermocellum CelJ is a modular enzyme containing a family 30 carbohydrate-binding module (CBM) and a family 9 catalytic module at its N-terminal moiety. To investigate the functions of the CBM and the catalytic module, truncated derivatives of CelJ were constructed and characterized. Isothermal titration calorimetric studies showed that the association constants (Ka ) of the CBM polypeptide (CBM30) for the binding of cellopentaose and cellohexaose were 1.2 × 104 and 6.4 × 104 M−1, respectively, and that the binding of CBM30 to these ligands is enthalpically driven. Qualitative analyses showed that CBM30 had strong affinity for cellulose and β-1,3-1,4-mixed glucan such as barley β-glucan and lichenan. Analyses of the hydrolytic action of the enzyme comprising the CBM and the catalytic module showed that the enzyme is a processive endoglucanse with strong activity towards carboxymethylcellulose, barley β-glucan and lichenan. By contrast, the catalytic module polypeptide devoid of the CBM showed negligible activity toward these substrates. These observations suggest that the CBM is extremely important not only because it mediates the binding of the enzyme to the substrates but also because it participates in the catalytic function of the enzyme or contributes to maintaining the correct tertiary structure of the family 9 catalytic module for expressing enzyme activity.

scholarly journals Novel Carbohydrate-Binding Module Identified in a Ruminal Metagenomic Endoglucanase

2010 ◽  
Vol 76 (14) ◽  
pp. 4867-4870 ◽  
Author(s):  
Cheng-Jie Duan ◽  
Jun-Liang Liu ◽  
Xi Wu ◽  
Ji-Liang Tang ◽  
Jia-Xun Feng
Keyword(s):  
Full Length ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
Carbohydrate Binding Modules ◽  
Significant Homology ◽  
Catalytic Module

ABSTRACT Endoglucanase C5614-1 comprises a catalytic module (CM) and an X module (XM). The XM showed no significant homology with known carbohydrate-binding modules (CBMs). Recombinant full-length endoglucanase could bind Avicel, whereas the CM could not. The XM could bind various polysaccharides. The results demonstrated that the XM was a new CBM.

scholarly journals Expression, purification and crystallization of a novel carbohydrate-binding module from theRuminococcus flavefacienscellulosome

2014 ◽  
Vol 70 (12) ◽  
pp. 1653-1656
Author(s):  
Immacolata Venditto ◽  
Maria S. J. Centeno ◽  
Luis M. A. Ferreira ◽  
Carlos M. G. A. Fontes ◽  
Shabir Najmudin
Keyword(s):  
Anaerobic Bacteria ◽  
Glycoside Hydrolase Family ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
The Novel ◽  
Carbohydrate Active Enzymes ◽  
Multiple Wavelength ◽  
Catalytic Module ◽  
Extensive Information ◽  
Hydrolase Family

Anaerobic bacteria organize carbohydrate-active enzymes into a multi-component complex, the cellulosome, which degrades cellulose and hemicellulose highly efficiently. Genome sequencing ofRuminococcus flavefaciensFD-1 offers extensive information on the range and diversity of the enzymatic and structural components of the cellulosome. TheR. flavefaciensFD-1 genome encodes over 200 dockerin-containing proteins, most of which are of unknown function. One of these modular proteins comprises a glycoside hydrolase family 5 catalytic module (GH5) linked to an unclassified carbohydrate-binding module (CBM-Rf1) and a dockerin. The novel CBM-Rf1 has been purified and crystallized. The crystals belonged to the trigonal space groupR32:H. The CBM-Rf1 structure was determined by a multiple-wavelength anomalous dispersion experiment usingAutoSolfrom thePHENIXsuite using both selenomethionyl-derivative and native data to resolutions of 2.28 and 2.0 Å, respectively.

scholarly journals Essentiality of a Newly Identified Carbohydrate-Binding Module for the Function of CelB (BH0603) from the Alkaliphilic Bacterium Bacillus halodurans

2006 ◽  
Vol 72 (10) ◽  
pp. 6851-6853 ◽  
Author(s):  
Benson Munyali Wamalwa ◽  
Makiko Sakka ◽  
Paul Mwanza Shiundu ◽  
Kunio Ohmiya ◽  
Tetsuya Kimura ◽  
...  
Keyword(s):  
Unknown Function ◽  
Glycoside Hydrolase ◽  
Bacillus Halodurans ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
Catalytic Module ◽  
Bacterium Bacillus ◽  
Alkaliphilic Bacterium

ABSTRACT CelB (BH0603) from Bacillus halodurans is a modular glycoside hydrolase with a family 5 catalytic module, an immunoglobulin-like module, and module PfamB of unknown function. The recombinant PfamB module bound to Avicel and was essential for CelB hydrolytic function. We propose that module PfamB be designated a new carbohydrate-binding module.

scholarly journals O-glycosylation effects on family 1 carbohydrate-binding module solution structures

FEBS Journal ◽  
2015 ◽  
Vol 282 (22) ◽  
pp. 4341-4356 ◽  
Author(s):  
Renee M. Happs ◽  
Xiaoyang Guan ◽  
Michael G. Resch ◽  
Mark F. Davis ◽  
Gregg T. Beckham ◽  
...  
Keyword(s):  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
Solution Structures

The N-terminal family 22 carbohydrate-binding module of xylanase 10B ofClostridium themocellumis not a thermostabilizing domain

2004 ◽  
Vol 238 (1) ◽  
pp. 71-78
Author(s):  
Fernando M.V. Dias ◽  
Arun Goyal ◽  
Harry J. Gilbert ◽  
José A.M. Prates ◽  
Luís M.A. Ferreira ◽  
...  
Keyword(s):  
Carbohydrate Binding ◽  
Carbohydrate Binding Module
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