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Current Proteomics

Editor-in-Chief

ISSN (Print): 1570-1646
ISSN (Online): 1875-6247

Research Article

Isolation, Identification and In Silico Study of Native Cellulase Producing Bacteria

Author(s): Farzane Kargar, Mojtaba Mortazavi*, Mahmood Maleki, Masoud Torkzadeh Mahani, Younes Ghasemi and Amir Savardashtaki*

Volume 18, Issue 1, 2021

Published on: 27 November, 2019

Page: [3 - 11] Pages: 9

DOI: 10.2174/1570164617666191127142035

Price: $65

Abstract

Aim: The purpose of this study was to screen the bacteria producing cellulase enzymes and their bioinformatics studies.

Background: Cellulose is a long-chain polymer of glucose that hydrolyzes by cellulases to glucose molecules. In order to design the new biotechnological applications, some strategies have been used as increasing the efficiency of enzyme production, generating cost-effective enzymes, producing stable enzymes and identification of new strains.

Objective: On the other hand, some bacteria special features have made them suitable candidates for the identification of the new source of enzymes. In this regard, some native strains of bacteria were screened.

Methods: These bacteria were grown on a culture containing the liquid M9 media containing CMC to ensure the synthesis of cellulase. The formation of a clear area in the culture medium indicated decomposition of cellulose. In the following, the DNA of these bacteria were extracted and their 16S rDNA genes were amplified.

Result: The results show that nine samples were able to synthesize cellulase. In following, these strains were identified using 16S rDNA. The results show that these screened bacteria belonged to the Bacillus sp., Alcaligenes sp., Alcaligenes sp., and Enterobacter sp.

Conclusion: The enzyme activity analysis shows that the Bacillus toyonensis, Bacillus sp. strain XA15-411 Bacillus cereus have produced the maximum yield of cellulases. However, these amounts of enzyme production in these samples are not proportional to their growth rate. As the bacterial growth chart within 4 consecutive days shows that the Alcaligenes sp. Bacillus cereus, Bacillus toyonensis, Bacillus sp. strain XA15-411 have a maximum growth rate. The study of the phylogenetic tree also shows that Bacillus species are more abundant in the production of cellulase enzyme. These bioinformatics analyses show that the Bacillus species have different evolutionary relationships and evolved in different evolutionary time. However, for maximum cellulase production by this bacteria, some information as optimum temperature, optimum pH, carbon and nitrogen sources are needed for the ideal formulation of media composition. The cellulase production is closely controlled in microorganisms and the cellulase yields appear to depend on a variety of factors. However, the further studies are needed for cloning, purification and application of these new microbial cellulases in the different commercial fields as in food, detergent, and pharmaceutical, paper, textile industries and also various chemical industries. However, these novel enzymes can be further engineered through rational design or using random mutagenesis techniques.

Keywords: 16S rDNA, Bacillus sp., cellulase, bioinformatics analysis, enzymes, biomass production.

Graphical Abstract

[1]
Paice, M.G.; Bernier, R., Jr; Jurasek, L. Viscosity-enhancing bleaching of hardwood kraft pulp with xylanase from a cloned gene. Biotechnol. Bioeng., 1988, 32(2), 235-239.
[http://dx.doi.org/10.1002/bit.260320214] [PMID: 18584740]
[2]
Gamerith, G.; Groicher, R.; Zeilinger, S.; Herzog, P.; Kubicek, C.P. Cellulase-poor xylanases produced by Trichoderma reesei RUT C-30 on hemicellulose substrates. Appl. Microbiol. Biotechnol., 1992, 38(3), 315-322.
[http://dx.doi.org/10.1007/BF00170079]
[3]
Li, W.; Zhang, W-W.; Yang, M-M.; Chen, Y-L. Cloning of the thermostable cellulase gene from newly isolated Bacillus subtilis and its expression in Escherichia coli. Mol. Biotechnol., 2008, 40(2), 195-201.
[http://dx.doi.org/10.1007/s12033-008-9079-y] [PMID: 18576142]
[4]
Miao, Y.; Li, J.; Xiao, Z.; Shen, Q.; Zhang, R. Characterization and identification of the xylanolytic enzymes from Aspergillus fumigatus Z5. BMC Microbiol., 2015, 15(1), 126.
[http://dx.doi.org/10.1186/s12866-015-0463-z] [PMID: 26100973]
[5]
Schülein, M. Protein engineering of cellulases. Biochimica et Biophysica Acta (BBA). Prot. Str. Mol. Enzymol., 2000, 1543(2), 239-252.
[http://dx.doi.org/10.1016/S0167-4838(00)00247-8]
[6]
Bayer, E.A.; Chanzy, H.; Lamed, R.; Shoham, Y. Cellulose, cellulases and cellulosomes. Curr. Opin. Struct. Biol., 1998, 8(5), 548-557.
[http://dx.doi.org/10.1016/S0959-440X(98)80143-7 ] [PMID: 9818257]
[7]
Grassick, A.; Murray, P.G.; Thompson, R.; Collins, C.M.; Byrnes, L.; Birrane, G.; Higgins, T.M.; Tuohy, M.G. Three-dimensional structure of a thermostable native cellobiohydrolase, CBH IB, and molecular characterization of the cel7 gene from the filamentous fungus, Talaromyces emersonii. Eur. J. Biochem., 2004, 271(22), 4495-4506.
[http://dx.doi.org/10.1111/j.1432-1033.2004.04409.x ] [PMID: 15560790]
[8]
Schwarz, W.H. The cellulosome and cellulose degradation by anaerobic bacteria. Appl. Microbiol. Biotechnol., 2001, 56(5-6), 634-649.
[http://dx.doi.org/10.1007/s002530100710] [PMID: 11601609]
[9]
Adlakha, N.; Sawant, S.; Anil, A.; Lali, A.; Yazdani, S.S. Specific fusion of β-1, 4-endoglucanase and β-1, 4-glucosidase enhances the cellulolytic activity and helps in channeling of the intermediates. Appl. Environ. Microbiol., 2012, 78(20), 01386-12.
[10]
Wilson, D.B.; Irwin, D.C. Genetics and properties of cellulases. Recent Progress in Bioconversion of Lignocellulosics; Springer, 1999, pp. 1-21.
[http://dx.doi.org/10.1007/3-540-49194-5_1]
[11]
Leschine, S.; Canale-Parola, E. Carbon cycling by cellulose-fermenting nitrogen-fixing bacteria. Adv. Space Res., 1989, 9(8), 149-152.
[http://dx.doi.org/10.1016/0273-1177(89)90039-2]
[12]
Lynd, L.R.; Weimer, P.J.; van Zyl, W.H.; Pretorius, I.S. Microbial cellulose utilization: Fundamentals and biotechnology. Microbiol. Mol. Biol. Rev., 2002, 66(3), 506-577.
[http://dx.doi.org/10.1128/MMBR.66.3.506-577.2002 ] [PMID: 12209002]
[13]
Pandey, S.; Kushwah, J.; Tiwari, R.; Kumar, R.; Somvanshi, V.S.; Nain, L.; Saxena, A.K. Cloning and expression of β-1, 4-endoglucanase gene from Bacillus subtilis isolated from soil long term irrigated with effluents of paper and pulp mill. Microbiol. Res., 2014, 169(9-10), 693-698.
[http://dx.doi.org/10.1016/j.micres.2014.02.006 ] [PMID: 24636744]
[14]
Helenius, G.; Bäckdahl, H.; Bodin, A.; Nannmark, U.; Gatenholm, P.; Risberg, B. In vivo biocompatibility of bacterial cellulose. J. Biomed. Mater. Res. A, 2006, 76(2), 431-438.
[http://dx.doi.org/10.1002/jbm.a.30570] [PMID: 16278860]
[15]
DEMİRBAŞ. A., Bioethanol from cellulosic materials: A renewable motor fuel from biomass. Energy Sources, 2005, 27(4), 327-337.
[http://dx.doi.org/10.1080/00908310390266643]
[16]
Kotchoni, O.d.; Shonukan, O.; Gachomo, W. Bacillus pumilus BpCRI 6, a promising candidate for cellulase production under conditions of catabolite repression. Afr. J. Biotechnol., 2003, 2(6), 140-146.
[http://dx.doi.org/10.5897/AJB2003.000-1028]
[17]
Feng, Y.; Duan, C-J.; Pang, H.; Mo, X-C.; Wu, C-F.; Yu, Y.; Hu, Y-L.; Wei, J.; Tang, J-L.; Feng, J-X. Cloning and identification of novel cellulase genes from uncultured microorganisms in rabbit cecum and characterization of the expressed cellulases. Appl. Microbiol. Biotechnol., 2007, 75(2), 319-328.
[http://dx.doi.org/10.1007/s00253-006-0820-9] [PMID: 17216439]
[18]
Galbe, M.; Zacchi, G. A review of the production of ethanol from softwood. Appl. Microbiol. Biotechnol., 2002, 59(6), 618-628.
[http://dx.doi.org/10.1007/s00253-002-1058-9] [PMID: 12226717]
[19]
Lee, Y-J.; Kim, B-K.; Lee, B-H.; Jo, K-I.; Lee, N-K.; Chung, C-H.; Lee, Y-C.; Lee, J-W. Purification and characterization of cellulase produced by Bacillus amyoliquefaciens DL-3 utilizing rice hull. Bioresour. Technol., 2008, 99(2), 378-386.
[http://dx.doi.org/10.1016/j.biortech.2006.12.013 ] [PMID: 17320379]
[20]
Mawadza, C.; Hatti-Kaul, R.; Zvauya, R.; Mattiasson, B. Purification and characterization of cellulases produced by two Bacillus strains. J. Biotechnol., 2000, 83(3), 177-187.
[http://dx.doi.org/10.1016/S0168-1656(00)00305-9 ] [PMID: 11051415]
[21]
Maki, M.; Leung, K.T.; Qin, W. The prospects of cellulase-producing bacteria for the bioconversion of lignocellulosic biomass. Int. J. Biol. Sci., 2009, 5(5), 500-516.
[http://dx.doi.org/10.7150/ijbs.5.500] [PMID: 19680472]
[22]
Juturu, V.; Wu, J.C. Microbial cellulases: Engineering, production and applications. Renew. Sustain. Energy Rev., 2014, 33, 188-203.
[http://dx.doi.org/10.1016/j.rser.2014.01.077]
[23]
Sadhu, S.; Maiti, T.K. Cellulase production by bacteria: A review. Br. Microbiol. Res. J., 2013, 3(3), 235.
[http://dx.doi.org/10.9734/BMRJ/2013/2367]
[24]
Kumar, S.; Stecher, G.; Tamura, K. MEGA7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol. Biol. Evol., 2016, 33(7), 1870-1874.
[http://dx.doi.org/10.1093/molbev/msw054] [PMID: 27004904]
[25]
Huson, D.H.; Bryant, D. User Manual for SplitsTree4., 2006, 4.
[26]
Miller, G.L. Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal. Chem., 1959, 31(3), 426-428.
[http://dx.doi.org/10.1021/ac60147a030]
[27]
Wood, T.M.; Bhat, K.M. Methods for measuring cellulase activities. Methods in enzymology; Elsevier, 1988, Vol. 160, pp. 87-112.
[28]
Suzuki, M.T.; Giovannoni, S.J. Bias caused by template annealing in the amplification of mixtures of 16S rRNA genes by PCR. Appl. Environ. Microbiol., 1996, 62(2), 625-630.
[PMID: 8593063]
[29]
Flandrois, J-P.; Perrière, G.; Gouy, M. leBIBIQBPP: A set of databases and a webtool for automatic phylogenetic analysis of prokaryotic sequences. BMC Bioinformatics, 2015, 16(1), 251.
[http://dx.doi.org/10.1186/s12859-015-0692-z] [PMID: 26264559]
[30]
Immanuel, G.; Dhanusha, R.; Prema, P.; Palavesam, A. Effect of different growth parameters on endoglucanase enzyme activity by bacteria isolated from coir retting effluents of estuarine environment. Int. J. Environ. Sci. Technol., 2006, 3(1), 25-34.
[http://dx.doi.org/10.1007/BF03325904]
[31]
Nakamura, K.; Kitamura, K. Isolation and identification of crystalline cellulose hydrolyzing bacterium and its enzymatic properties. J. Ferment. Technol., 1982, 60(4), 343-348.
[32]
Kargar, F.; Mortazavi, M.; Savardashtaki, A.; Hosseinkhani, S.; Mahani, M.T.; Ghasemi, Y. Genomic and protein structure analysis of the luciferase from the Iranian bioluminescent beetle, Luciola sp. Int. J. Biol. Macromol., 2019, 124, 689-698.
[http://dx.doi.org/10.1016/j.ijbiomac.2018.11.264 ] [PMID: 30502432]
[33]
Mortazavi, M.; Hosseinkhani, S. Surface charge modification increases firefly luciferase rigidity without alteration in bioluminescence spectra. Enzyme Microb. Technol., 2017, 96, 47-59.
[http://dx.doi.org/10.1016/j.enzmictec.2016.09.014 ] [PMID: 27871385]
[34]
Yousefi, F.; Ataei, F.; Mortazavi, M.; Hosseinkhani, S. Bifunctional role of leucine 300 of firefly luciferase in structural rigidity. Int. J. Biol. Macromol., 2017, 101, 67-74.
[http://dx.doi.org/10.1016/j.ijbiomac.2017.03.069 ] [PMID: 28322945]
[35]
Mortazavi, M.; Nezafat, N.; Negahdaripour, M.; Gholami, A.; Torkzadeh-Mahani, M.; Lotfi, S.; Ghasemi, Y. In silico evaluation of rare codons and their positions in the structure of cytosine deaminase and substrate docking studies. Trends Pharmacol. Sci., 2016, 2(2)
[36]
Mortazavi, M.; Zarenezhad, M.; Gholamzadeh, S.; Alavian, S.M.; Ghorbani, M.; Dehghani, R.; Malekpour, A.; Meshkibaf, M.; Fakhrzad, A. Bioinformatic identification of Rare Codon Clusters (RCCs) in HBV genome and evaluation of RCCs in proteins structure of hepatitis B virus. Hepat. Mon., 2016, 16(10)e39909
[http://dx.doi.org/10.5812/hepatmon.39909] [PMID: 27882067]
[37]
Sethi, S.; Datta, A.; Gupta, B.L.; Gupta, S. Optimization of cellulase production from bacteria isolated from soil. ISRN biotechnology., 2013, 2013.
[http://dx.doi.org/10.5402/2013/985685]
[38]
Bajaj, B.K.; Singh, N.P. Production of xylanase from an alkali tolerant Streptomyces sp. 7b under solid-state fermentation, its purification, and characterization. Appl. Biochem. Biotechnol., 2010, 162(6), 1804-1818.
[http://dx.doi.org/10.1007/s12010-010-8960-x] [PMID: 20419509]
[39]
Mahanta, N.; Gupta, A.; Khare, S.K. Production of protease and lipase by solvent tolerant Pseudomonas aeruginosa PseA in solid-state fermentation using Jatropha curcas seed cake as substrate. Bioresour. Technol., 2008, 99(6), 1729-1735.
[http://dx.doi.org/10.1016/j.biortech.2007.03.046 ] [PMID: 17509877]
[40]
Dhawan, S.; Kaur, J. Microbial mannanases: An overview of production and applications. Crit. Rev. Biotechnol., 2007, 27(4), 197-216.
[http://dx.doi.org/10.1080/07388550701775919 ] [PMID: 18085462]

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