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Molecular Characterization of a Thermophilic and Salt- and Alkaline-Tolerant Xylanase from Planococcus sp. SL4, a Strain Isolated from the Sediment of a Soda Lake
1College of Biological Science and Technology, Fuzhou University, Fuzhou 350108, P.R. China, 2Fujian Key Laboratory of Marine Enzyme Engineering, Fuzhou 350002, P.R. China
J. Microbiol. Biotechnol. 2015; 25(5): 662-671
Published May 28, 2015 https://doi.org/10.4014/jmb.1408.08062
Copyright © The Korean Society for Microbiology and Biotechnology.
Abstract
Keywords
References
- Anbarasan S, Janis J, Paloheimo M, Laitaoja M, Vuolanto M, Karimaki J, et al. 2010. Effect of glycosylation and additional domains on the thermostability of a family 10 xylanase produced by Thermopolyspora flexuosa. Appl. Environ. Microbiol. 76: 356-360.
- Antony CP, Kumaresan D, Hunger S, Drake HL, Murrell JC, Shouche YS. 2013. Microbiology of Lonar Lake and other soda lakes. ISME J. 7: 468-476.
- Bai W, Xue Y, Zhou C, Ma Y. 2012. Cloning, expression and characterization of a novel salt-tolerant xylanase from Bacillus sp. SN5. Biotechnol. Lett. 34: 2093-2099.
- Bastawde KB. 1992. Xylan structure, microbial xylanases, and their mode of action. World J. Microbiol. Biotechnol. 8:353-368.
- Beg QK, Kapoor M, Mahajan L, Hoondal GS. 2001. Microbial xylanases and their industrial applications: a review. Appl. Microbiol. Biotechnol. 56: 326-338.
- Biely P, Vrsanska M, Tenkanen M, Kluepfel D. 1997. Endoβ-1,4-xylanase families: differences in catalytic properties. J. Biotechnol. 57: 151-166.
- Bradford MM. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248-254.
- Cantarel BL, Coutinho PM, Rancurel C, Bernard T, Lombard V, Henrissat B. 2009. The carbohydrate-active enzymes database (CAZy): an expert resource for glycogenomics. Nucleic Acids Res. 37: D233-D238.
- Chang P, Tsai W S, T sai CL, Tseng MJ. 2 004. C loning and characterization of two thermostable xylanases from an alkaliphilic Bacillus firmus. Biochem. Biophys. Res. Commun. 319: 1017-1025.
- Collins T, Gerday C, Feller G. 2005. Xylanases, xylanase families and extremophilic xylanases. FEMS Microbiol. Rev. 29: 3-23.
- Ferreira-Filho EX. 1994. The xylan-degrading enzyme system. Braz. J. Med. Biol. Res. 27: 1093-1109.
- Fukuchi S, Yoshimune K, Wakayama M, Moriguchi M, Nishikawa K. 2003. Unique amino acid composition of proteins in halophilic bacteria. J. Mol. Biol. 327: 347-357.
- Grant W, Sorokin D. 2011. Distribution and diversity of soda lake alkaliphiles, pp. 27-54. In Horikoshi K (ed.). Extremophiles Handbook. Springer, Japan.
- Guo B, Chen XL, Sun CY, Zhou BC, Zhang YZ. 2009. Gene cloning, expression and characterization of a new coldactive and salt-tolerant endo-β-1,4-xylanase from marine Glaciecola mesophila KMM 241. Appl. Microbiol. Biotechnol. 84:1107-1115.
- Gupta N, Reddy VS, Maiti S, Ghosh A. 2000. Cloning, expression, and sequence analysis of the gene encoding the alkali-stable, thermostable endoxylanase from alkalophilic, mesophilic Bacillus sp. strain NG-27. Appl. Environ. Microbiol. 66: 2631-2635.
- Hung KS, L iu SM, F ang TY, T zou WS, Lin FP, Sun KH, Tang SJ. 2011. Characterization of a salt-tolerant xylanase from Thermoanaerobacterium saccharolyticum NTOU1. Biotechnol. Lett. 33: 1441-1447.
- Ito S. 2011. Alkaline enzymes in current detergency, pp. 229-251. In Horikoshi K (ed.). Extremophiles Handbook. Springer, Japan.
- Jeffries TW. 1996. Biochemistry and genetics of microbial xylanases. Curr. Opin. Biotechnol. 7: 337-342.
- Jones BE, Grant WD, Duckworth AW, Owenson GG. 1998. Microbial diversity of soda lakes. Extremophiles 2: 191-200.
- Kamal Kumar B, Balakrishnan H, Rele MV. 2004. Compatibility of alkaline xylanases from an alkaliphilic Bacillus NCL (87-6-10) with commercial detergents and proteases. J. Ind. Microbiol. Biotechnol. 31: 83-87.
- Khasin A, Alchanati I, Shoham Y. 1993. Purification and characterization of a thermostable xylanase from Bacillus stearothermophilus T-6. Appl. Environ. Microbiol. 59: 1725-1730.
- Kulkarni N, Shendye A, Rao M. 1999. Molecular and biotechnological aspects of xylanases. FEMS Microbiol. Rev. 23: 411-456.
- Liu B, Zhang N, Zhao C, Lin B, Xie L, Huang Y. 2012. Characterization of a recombinant thermostable xylanase from hot spring thermophilic Geobacillus sp. TC-W7. J. Microbiol. Biotechnol. 22: 1388-1394.
- Liu X, Huang Z, Zhang X, Shao Z, Liu Z. 2014. Cloning, expression and characterization of a novel cold-active and halophilic xylanase from Zunongwangia profunda. Extremophiles 18: 441-450.
- Liu Y-G, Whittier RF. 1995. Thermal asymmetric interlaced PCR: automatable amplification and sequencing of insert end fragments from P1 and YAC clones for chromosome walking. Genomics 25: 674-681.
- Mamo G, Delgado O, Martinez A, Mattiasson B, Hatti-Kaul R. 2006. Cloning, sequence analysis, and expression of a gene encoding an endoxylanase from Bacillus halodurans S7. Mol. Biotechnol. 33: 149-159.
- Mamo G, Thunnissen M, Hatti-Kaul R, Mattiasson B. 2009. An alkaline active xylanase: insights into mechanisms of high pH catalytic adaptation. Biochimie 91: 1187-1196.
- Manikandan K, Bhardwaj A, Gupta N, Lokanath NK, Ghosh A, Reddy VS, Ramakumar S. 2006. Crystal structures of native and xylosaccharide-bound alkali thermostable xylanase from an alkalophilic Bacillus sp. NG-27: structural insights into alkalophilicity and implications for adaptation to polyextreme conditions. Protein Sci. 15: 1951-1960.
- Mielenz JR. 2001. Ethanol production from biomass: technology and commercialization status. Curr. Opin. Microbiol. 4: 324-329.
- Miller GL, Blum R, Glennon WE, Burton AL. 1960. Measurement of carboxymethylcellulase activity. Anal. Biochem. 1: 127-132.
- Nimchua T, Thongaram T, Uengwetwanit T, Pongpattanakitshote S, Eurwilaichitr L. 2012. Metagenomic analysis of novel lignocellulose-degrading enzymes from higher termite guts inhabiting microbes. J. Microbiol. Biotechnol. 22: 462-469.
- Petersen TN, Brunak S, von Heijne G, Nielsen H. 2011. SignalP 4.0: discriminating signal peptides from transmembrane regions. Nat. Methods 8: 785-786.
- Prade RA. 1996. Xylanases: from biology to biotechnology. Biotechnol. Genet. Eng. Rev. 13: 101-131.
- Saha BC. 2003. Hemicellulose bioconversion. J. Ind. Microbiol. Biotechnol. 30: 279-291.
- Shen J, Cao JT, Wu YH. 2001. Paleoclimatic changes in Dabusu Lake. Chin. J. Oceanol. Limnol. 19: 91-96
- Simkhada JR, Yoo HY, Choi YH, Kim SW, Yoo JC. 2012. An extremely alkaline novel xylanase from a newly isolated Streptomyces strain cultivated in corncob medium. Appl. Biochem. Biotechnol. 168: 2017-2027.
- Tamura K, Dudley J, Nei M, Kumar S. 2007. MEGA4:molecular evolutionary genetics analysis (MEGA) software version 4.0. Mol. Biol. Evol. 24: 1596-1599.
- van den Burg B. 2003. Extremophiles as a source for novel enzymes. Curr. Opin. Microbiol. 6: 213-218.
- Verma D, Kawarabayasi Y, Miyazaki K, Satyanarayana T. 2013. Cloning, expression and characteristics of a novel alkalistable and thermostable xylanase encoding gene (Mxyl) retrieved from compost-soil metagenome. PLoS One 8: e52459.
- Verma D, Satyanarayana T. 2012. Cloning, expression and applicability of thermo-alkali-stable xylanase of Geobacillus thermoleovorans in generating xylooligosaccharides from agro-residues. Bioresour. Technol. 107: 333-338.
- Viikari L, Kantelinen A, Sundquist J, Linko M. 1994. Xylanases in bleaching: from an idea to the industry. FEMS Microbiol. Rev. 13: 335-350.
- Wang G, Wang Y, Yang P, Luo H, Huang H, Shi P, et al. 2010. Molecular detection and diversity of xylanase genes in alpine tundra soil. Appl. Microbiol. Biotechnol. 87: 1383-1393.
- Xiong H, Nyyssölä A, Jänis J, Pastinen O, Weymarn Nv, Leisola M, Turunen O. 2004. Characterization of the xylanase produced by submerged cultivation of Thermomyces lanuginosus DSM 10635. Enzyme Microb. Technol. 35: 93-99.
- Zhang G, Mao L, Zhao Y, Xue Y, Ma Y. 2010. Characterization of a thermostable xylanase from an alkaliphilic Bacillus sp. Biotechnol. Lett. 32: 1915-1920.
- Zhao Y, Luo H, Meng K, Shi P, Wang G, Yang P, et al. 2011. A xylanase gene directly cloned from the genomic DNA of alkaline wastewater sludge showing application potential in the paper industry. Appl. Biochem. Biotechnol. 165: 35-46.
- Zhao Y, M eng K, Luo H, Y ang P, Shi P, Huang H, et al. 2011. Cloning, expression, and characterization of a new xylanase from alkalophilic Paenibacillus sp. 12-11. J. Microbiol. Biotechnol. 21: 861-868.
- Zhou J, Gao Y, Dong Y, Tang X, Li J, Xu B, et al. 2012. A novel xylanase with tolerance to ethanol, salt, protease, SDS, heat, and alkali from actinomycete Lechevalieria sp. HJ3. J. Ind. Microbiol. Biotechnol. 39: 965-975.
Related articles in JMB

Article
Research article
J. Microbiol. Biotechnol. 2015; 25(5): 662-671
Published online May 28, 2015 https://doi.org/10.4014/jmb.1408.08062
Copyright © The Korean Society for Microbiology and Biotechnology.
Molecular Characterization of a Thermophilic and Salt- and Alkaline-Tolerant Xylanase from Planococcus sp. SL4, a Strain Isolated from the Sediment of a Soda Lake
Xiaoyun Huang 1, Juan Lin 1, 2, Xiuyun Ye 1, 2 and Guozeng Wang 1, 2*
1College of Biological Science and Technology, Fuzhou University, Fuzhou 350108, P.R. China, 2Fujian Key Laboratory of Marine Enzyme Engineering, Fuzhou 350002, P.R. China
Abstract
To enrich the genetic resource of microbial xylanases with high activity and stability under
alkaline conditions, a xylanase gene (xynSL4) was cloned from Planococcus sp. SL4, an alkaline
xylanase-producing strain isolated from the sediment of soda lake Dabusu. Deduced XynSL4
consists of a putative signal peptide of 29 residues and a catalytic domain (30-380 residues) of
glycosyl hydrolase family 10, and shares the highest identity of 77% with a hypothetical
protein from Planomicrobium glaciei CHR43. Phylogenetic analysis indicated that deduced
XynSL4 is closely related with thermophilic and alkaline xylanases from Geobacillus and
Bacillus species. The gene xynSL4 was expressed heterologously in Escherichia coli and the
recombinant enzyme showed some superior properties. Purified recombinant XynSL4
(rXynSL4) was highly active and stable over the neutral and alkaline pH range from 6 to 11,
with maximum activity at pH 7 and more than 60% activity at pH 11. It had an apparent
temperature optimum of 70oC and retained stable at this temperature in the presence of
substrate. rXynSL4 was highly halotolerant, retaining more than 55% activity with 0.25–3.0 M
NaCl and was stable at the concentration of NaCl up to 4M. The enzyme activity was
significantly enhanced by β-mercaptoethanol and Ca2+ but strongly inhibited by heavy-metal
ions and SDS. This thermophilic and alkaline- and salt-tolerant enzyme has great potential for
basic research and industrial applications.
Keywords: Xylanase, Planococcus, Gene cloning, Thermophilic, Alkaline and salt-tolerant
References
- Anbarasan S, Janis J, Paloheimo M, Laitaoja M, Vuolanto M, Karimaki J, et al. 2010. Effect of glycosylation and additional domains on the thermostability of a family 10 xylanase produced by Thermopolyspora flexuosa. Appl. Environ. Microbiol. 76: 356-360.
- Antony CP, Kumaresan D, Hunger S, Drake HL, Murrell JC, Shouche YS. 2013. Microbiology of Lonar Lake and other soda lakes. ISME J. 7: 468-476.
- Bai W, Xue Y, Zhou C, Ma Y. 2012. Cloning, expression and characterization of a novel salt-tolerant xylanase from Bacillus sp. SN5. Biotechnol. Lett. 34: 2093-2099.
- Bastawde KB. 1992. Xylan structure, microbial xylanases, and their mode of action. World J. Microbiol. Biotechnol. 8:353-368.
- Beg QK, Kapoor M, Mahajan L, Hoondal GS. 2001. Microbial xylanases and their industrial applications: a review. Appl. Microbiol. Biotechnol. 56: 326-338.
- Biely P, Vrsanska M, Tenkanen M, Kluepfel D. 1997. Endoβ-1,4-xylanase families: differences in catalytic properties. J. Biotechnol. 57: 151-166.
- Bradford MM. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248-254.
- Cantarel BL, Coutinho PM, Rancurel C, Bernard T, Lombard V, Henrissat B. 2009. The carbohydrate-active enzymes database (CAZy): an expert resource for glycogenomics. Nucleic Acids Res. 37: D233-D238.
- Chang P, Tsai W S, T sai CL, Tseng MJ. 2 004. C loning and characterization of two thermostable xylanases from an alkaliphilic Bacillus firmus. Biochem. Biophys. Res. Commun. 319: 1017-1025.
- Collins T, Gerday C, Feller G. 2005. Xylanases, xylanase families and extremophilic xylanases. FEMS Microbiol. Rev. 29: 3-23.
- Ferreira-Filho EX. 1994. The xylan-degrading enzyme system. Braz. J. Med. Biol. Res. 27: 1093-1109.
- Fukuchi S, Yoshimune K, Wakayama M, Moriguchi M, Nishikawa K. 2003. Unique amino acid composition of proteins in halophilic bacteria. J. Mol. Biol. 327: 347-357.
- Grant W, Sorokin D. 2011. Distribution and diversity of soda lake alkaliphiles, pp. 27-54. In Horikoshi K (ed.). Extremophiles Handbook. Springer, Japan.
- Guo B, Chen XL, Sun CY, Zhou BC, Zhang YZ. 2009. Gene cloning, expression and characterization of a new coldactive and salt-tolerant endo-β-1,4-xylanase from marine Glaciecola mesophila KMM 241. Appl. Microbiol. Biotechnol. 84:1107-1115.
- Gupta N, Reddy VS, Maiti S, Ghosh A. 2000. Cloning, expression, and sequence analysis of the gene encoding the alkali-stable, thermostable endoxylanase from alkalophilic, mesophilic Bacillus sp. strain NG-27. Appl. Environ. Microbiol. 66: 2631-2635.
- Hung KS, L iu SM, F ang TY, T zou WS, Lin FP, Sun KH, Tang SJ. 2011. Characterization of a salt-tolerant xylanase from Thermoanaerobacterium saccharolyticum NTOU1. Biotechnol. Lett. 33: 1441-1447.
- Ito S. 2011. Alkaline enzymes in current detergency, pp. 229-251. In Horikoshi K (ed.). Extremophiles Handbook. Springer, Japan.
- Jeffries TW. 1996. Biochemistry and genetics of microbial xylanases. Curr. Opin. Biotechnol. 7: 337-342.
- Jones BE, Grant WD, Duckworth AW, Owenson GG. 1998. Microbial diversity of soda lakes. Extremophiles 2: 191-200.
- Kamal Kumar B, Balakrishnan H, Rele MV. 2004. Compatibility of alkaline xylanases from an alkaliphilic Bacillus NCL (87-6-10) with commercial detergents and proteases. J. Ind. Microbiol. Biotechnol. 31: 83-87.
- Khasin A, Alchanati I, Shoham Y. 1993. Purification and characterization of a thermostable xylanase from Bacillus stearothermophilus T-6. Appl. Environ. Microbiol. 59: 1725-1730.
- Kulkarni N, Shendye A, Rao M. 1999. Molecular and biotechnological aspects of xylanases. FEMS Microbiol. Rev. 23: 411-456.
- Liu B, Zhang N, Zhao C, Lin B, Xie L, Huang Y. 2012. Characterization of a recombinant thermostable xylanase from hot spring thermophilic Geobacillus sp. TC-W7. J. Microbiol. Biotechnol. 22: 1388-1394.
- Liu X, Huang Z, Zhang X, Shao Z, Liu Z. 2014. Cloning, expression and characterization of a novel cold-active and halophilic xylanase from Zunongwangia profunda. Extremophiles 18: 441-450.
- Liu Y-G, Whittier RF. 1995. Thermal asymmetric interlaced PCR: automatable amplification and sequencing of insert end fragments from P1 and YAC clones for chromosome walking. Genomics 25: 674-681.
- Mamo G, Delgado O, Martinez A, Mattiasson B, Hatti-Kaul R. 2006. Cloning, sequence analysis, and expression of a gene encoding an endoxylanase from Bacillus halodurans S7. Mol. Biotechnol. 33: 149-159.
- Mamo G, Thunnissen M, Hatti-Kaul R, Mattiasson B. 2009. An alkaline active xylanase: insights into mechanisms of high pH catalytic adaptation. Biochimie 91: 1187-1196.
- Manikandan K, Bhardwaj A, Gupta N, Lokanath NK, Ghosh A, Reddy VS, Ramakumar S. 2006. Crystal structures of native and xylosaccharide-bound alkali thermostable xylanase from an alkalophilic Bacillus sp. NG-27: structural insights into alkalophilicity and implications for adaptation to polyextreme conditions. Protein Sci. 15: 1951-1960.
- Mielenz JR. 2001. Ethanol production from biomass: technology and commercialization status. Curr. Opin. Microbiol. 4: 324-329.
- Miller GL, Blum R, Glennon WE, Burton AL. 1960. Measurement of carboxymethylcellulase activity. Anal. Biochem. 1: 127-132.
- Nimchua T, Thongaram T, Uengwetwanit T, Pongpattanakitshote S, Eurwilaichitr L. 2012. Metagenomic analysis of novel lignocellulose-degrading enzymes from higher termite guts inhabiting microbes. J. Microbiol. Biotechnol. 22: 462-469.
- Petersen TN, Brunak S, von Heijne G, Nielsen H. 2011. SignalP 4.0: discriminating signal peptides from transmembrane regions. Nat. Methods 8: 785-786.
- Prade RA. 1996. Xylanases: from biology to biotechnology. Biotechnol. Genet. Eng. Rev. 13: 101-131.
- Saha BC. 2003. Hemicellulose bioconversion. J. Ind. Microbiol. Biotechnol. 30: 279-291.
- Shen J, Cao JT, Wu YH. 2001. Paleoclimatic changes in Dabusu Lake. Chin. J. Oceanol. Limnol. 19: 91-96
- Simkhada JR, Yoo HY, Choi YH, Kim SW, Yoo JC. 2012. An extremely alkaline novel xylanase from a newly isolated Streptomyces strain cultivated in corncob medium. Appl. Biochem. Biotechnol. 168: 2017-2027.
- Tamura K, Dudley J, Nei M, Kumar S. 2007. MEGA4:molecular evolutionary genetics analysis (MEGA) software version 4.0. Mol. Biol. Evol. 24: 1596-1599.
- van den Burg B. 2003. Extremophiles as a source for novel enzymes. Curr. Opin. Microbiol. 6: 213-218.
- Verma D, Kawarabayasi Y, Miyazaki K, Satyanarayana T. 2013. Cloning, expression and characteristics of a novel alkalistable and thermostable xylanase encoding gene (Mxyl) retrieved from compost-soil metagenome. PLoS One 8: e52459.
- Verma D, Satyanarayana T. 2012. Cloning, expression and applicability of thermo-alkali-stable xylanase of Geobacillus thermoleovorans in generating xylooligosaccharides from agro-residues. Bioresour. Technol. 107: 333-338.
- Viikari L, Kantelinen A, Sundquist J, Linko M. 1994. Xylanases in bleaching: from an idea to the industry. FEMS Microbiol. Rev. 13: 335-350.
- Wang G, Wang Y, Yang P, Luo H, Huang H, Shi P, et al. 2010. Molecular detection and diversity of xylanase genes in alpine tundra soil. Appl. Microbiol. Biotechnol. 87: 1383-1393.
- Xiong H, Nyyssölä A, Jänis J, Pastinen O, Weymarn Nv, Leisola M, Turunen O. 2004. Characterization of the xylanase produced by submerged cultivation of Thermomyces lanuginosus DSM 10635. Enzyme Microb. Technol. 35: 93-99.
- Zhang G, Mao L, Zhao Y, Xue Y, Ma Y. 2010. Characterization of a thermostable xylanase from an alkaliphilic Bacillus sp. Biotechnol. Lett. 32: 1915-1920.
- Zhao Y, Luo H, Meng K, Shi P, Wang G, Yang P, et al. 2011. A xylanase gene directly cloned from the genomic DNA of alkaline wastewater sludge showing application potential in the paper industry. Appl. Biochem. Biotechnol. 165: 35-46.
- Zhao Y, M eng K, Luo H, Y ang P, Shi P, Huang H, et al. 2011. Cloning, expression, and characterization of a new xylanase from alkalophilic Paenibacillus sp. 12-11. J. Microbiol. Biotechnol. 21: 861-868.
- Zhou J, Gao Y, Dong Y, Tang X, Li J, Xu B, et al. 2012. A novel xylanase with tolerance to ethanol, salt, protease, SDS, heat, and alkali from actinomycete Lechevalieria sp. HJ3. J. Ind. Microbiol. Biotechnol. 39: 965-975.