A Comparative Study of Activity and Stability of the Free and the Immobilized Endoglucanase from Alicyclobacillus Acidocaldarius

Document Type: Article

Authors

1 Department of Cellular and Molecular Biology, Faculty of Science, Azarbaijan Shahid Madani University, Tabriz, Iran.

2 Department of Cellular and Molecular Biology, Faculty of Sciences, Azarbaijan Shahid Madani University, Tabriz, Iran

Abstract

AaCel9A [β-1,4-endoglucanase, (E.C:3.2.1.4)], was immobilized onto glutaraldehyde activated chitosan macrosphere by covalent attachment. The properties of the immobilized AaCel9A were investigated by determining the optimum pH and optimum temperature for activity, thermal stability, and kinetic parameters. The immobilization process shifted the enzyme’s optimum temperature from 65 °C for the free enzyme towards a wider temperature range from 60-80 °C by the immobilized enzyme. The optimum pH of immobilized AaCel9A shifted to basic pH (pH 8) relative to free AaCel9A (pH 6.5). The immobilization on chitosan macrosphere enhanced half-life of AaCel9A enzyme. After 60 min, the immobilized and the free enzyme retained 75% and 40% their activity at 65 °C, respectively. The immobilized enzyme showed higher thermal stability than the free form. Km value of immobilized AaCel9A (17.05 mg ml-1) was higher than free AaCel9A (7.75 mg ml-1). Also, CMC hydrolysis by immobilized and free AaCel9A in the presence of SDS detergent was investigated. The results showed that the immobilized enzyme maintained its activity more than the free form in different concentrations of SDS.

Graphical Abstract

A Comparative Study of Activity and Stability of the Free and the Immobilized Endoglucanase from Alicyclobacillus Acidocaldarius

Keywords

Main Subjects


 

[1]         S. Acharya, A. Chaudhary, Braz. J. Microbiol. 43 (2012) 844.

[2]         W.M. Fogarty, C.T. Kelly, Microbial Enzymes and Biotechnology, Springer Science & Business Media, 2012.

[3]         Y. Cao, H. Tan, Carbohyd. Res. 337 (2002) 1291.

[4]         T.V. Vuong, D.B. Wilson, Biotechnol. Bioeng. 107 (2010) 195.

[5]         K. Komarova, Strategy For Cellulase Immobilization and its Partial Purification and Characterization, 2008.

[6]         K. Eckert, F. Zielinski, L.L. Leggio, E. Schneider, Appl. Microbiol. Biot. 60 (2002) 428.

[7]         K. Eckert, A. Vigouroux, L.L. Leggio, S. Moréra, J. Mol. Biol. 394 (2009) 61.

[8]         H. Liu, J.H. Pereira, P.D. Adams, R. Sapra, B.A. Simmons, K.L. Sale, FEBS lett. 584 (2010) 3431.

[9]         J.H. Pereira, R. Sapra, J.V. Volponi, C.L. Kozina, B. Simmons, P.D. Adams, Acta Crystallogr. D: Biol. Crystallogr. 65 (2009) 744.

[10]     R.C. Kuhad, R. Gupta, A. Singh, Enzyme Res. 2011 (2011).

[11]     S. Sadhu, T.K. Maiti, Brit. Microbiol. Res. J. 3 (2013) 235.

[12]     N. Sarkar, S.K. Ghosh, S. Bannerjee, K. Aikat, Renew. Energ. 37 (2012) 19.

[13]     H. Liu, P.H. Jose , P.D. Adams, R. Sapra, B.A. Simmons, K.L. Sale, FEBS Lett. 584 (2010) 3431.

[14]     V.B. Agbor, N. Cicek, R. Sparling, A. Berlin, D.B. Levin, Biotechnol. Adv. 29 (2011) 675.

[15]     L. Viikari, J. Vehmaanperä, A. Koivula, Biomass Bioenerg. 46 (2012) 13.

[16]     P.V. Iyer, L. Ananthanarayan, Process Biochem. 43 (2008) 1019.

[17]     V. Stepankova, S. Bidmanova, T. Koudelakova, Z. Prokop, R. Chaloupkova, J. Damborsky, Acs Catal. 3 (2013) 2823.

[18]     C. Mateo, J.M. Palomo, G. Fernandez-Lorente, J.M. Guisan, R. Enzyme Microb. Tech. 40 (2007) 1451.

[19]     M. Kamburov, I. Lalov, Biotechnol. Biotechnological Equipment 26 (2012) 156.

[20]     E. Górecka, M. Jastrzębska, Biotechnol. Food Sci. 75 (2011) 65.

[21]     S. Datta, L.R. Christena, Y.R.S. Rajaram, Biotech. 3 (2013) 1.

[22]     H. Chen, Q. Zhang, Y. Dang, G. Shu, Adv. J. Food Sci. Technol. 5 (2013) 932.

[23]     B. Krajewska, Enzyme Microb. Tech. 35 (2004) 126.

[24]     A. Rampino, M. Borgogna, P. Blasi, B. Bellich, A. Cesàro, Int. J. Pharm. 455 (2013) 219.

[25]     F.S. Younesi, M. Pazhang, S. Najavand, P. Rahimizadeh, M. Akbarian, M. Mohammadian, K. Khajeh, Mol. Biotechnol. 58 (2016) 12.

[26]     G.L. Miller, Anal. Chem. 31 (1959) 426.

[27]     E. Biró, Á.S. Németh, C. Sisak, T. Feczkó, J. Gyenis, J. Biochem. Bioph. Meth. 70 (2008) 1240.

[28]     K. Belho, S. Nongpiur, P. Ambasht, J. Protein. Proteomics 5 (2014) 177.

[29]     M.M. Bradford, Anal. Biochem. 72 (1976) 248.

[30]     A.A. Homaei, R. Sariri, F. Vianello, R. Stevanato, J. Chem. Biol. 6 (2013) 185.

[31]     P.V. Iyer, L. Ananthanarayan, Process Biochem. 43 (2008) 1019.

[32]     N.R. Mohamad, N.H.C. Marzuki, N.A. Buang, F. Huyop, R.A. Wahab, Biotechnol. Biotechnological Equipment 29 (2015) 205.

[33]     A. Suescun, N. Rueda, J.C. dos Santos, J.J. Castillo, C. Ortiz, R. Torres, O. Barbosa, R. Fernandez-Lafuente, Process Biochem. 50 (2015) 1211.

[34]     P.K. Srivastava, A. Anand, Int. J. Biol. Macromol. 64 (2014) 150.

[35]     N. Singh, A.M. Kayastha, Carbohyd. Res. 358 (2012) 61.

[36]     S.H. Chiou, T.C. Hung, R. Giridhar, W.T. Wu, Prep. Biochem. Biotech. 37 (2007) 265.

[37]     P. Tripathi, A. Kumari, P. Rath, A.M. Kayastha, Journal of Molecular Catalysis B: Enzymatic, 49 (2007) 69.

[38]     A. Kumari, A.M. Kayastha, J. Mol. Catal. B-Enzym. 69 (2011) 8.

[39]     M.A. El-Ghaffar, M. Hashem, Carbohyd. Polym. 81 (2010) 507.

[40]     F. Jafary, J. Varshosaz, M. Panjehpour, P. Yaghmaei, Russ. J. Appl. Chem. 87 (2014) 1719.

[41]     M. Kamburov, I. Lalov, Pharmaceutical Biotechnol. 26 (2011) 156.

[42]     A.N. Singh, S. Singh, N. Suthar, V.K. Dubey, J. Agr. Food Chem. 59 (2011) 6256.

[43]     S.A.S. Çetinus, H.N. Öztop, Enzyme Microb. Tech. 32 (2003) 889.

[44]     S. Moh’d A, J. Wiegel, Open Biochem. J. 4 (2010) 22.

[45]S. Ahmed, N. El-Shayeb, A. Hashem, S. Saleh, A. Abdel-Fattah, Braz. J. Chem. Eng. 30 (2013) 747.