Abstract
The β-glucosidase gene, bglC, was cloned from Bacillus sp. SJ-10 isolated from the squid jeotgal. Recombinant BglC protein overexpression was induced in Escherichia coli. The optimal pH and temperature of the enzyme, using p-nitrophenyl-β-d-glucopyranoside (pNPβGlc) as a substrate, were pH 6 and 40 °C, respectively. Enzymatic activity increased by 3.3- and 3.5-fold in the presence of 15 % NaCl and KCl, respectively. Furthermore, enzyme thermostability improved in the presence of NaCl or KCl. At 45 °C in the presence of salts, the enzyme was stable for 2 h and maintained 80 % activity. In the absence of salts, BglC completely lost activity after 110 min at 45 °C. Comparison of the kinetic parameters at various salt concentrations revealed that BglC had approximately 1.5- and 1.2-fold higher affinity and hydrolyzed pNPβGlc 1.9- and 2.1-fold faster in the presence of 15 % NaCl and KCl, respectively. Additionally, the Gibb’s free energy for denaturation was higher in the presence of 15 % salt than in the absence of salt at 45 and 50 °C. Since enzymatic activity and thermostability were enhanced under high salinity conditions, BglC is an ideal salt-tolerant enzyme for further research and industrial applications.
Similar content being viewed by others
References
Ketudat Cairns JR, Esen A (2010) β-Glucosidases. Cell Mol Life Sci 67:3389–3405
Singhania RR, Patel AK, Sukumaran RK, Larroche C, Pandey A (2012) Role and significance of beta-glucosidases in the hydrolysis of cellulose for bioethanol production. Bioresource Technol 127:500–507
Bhatia Y, Mishra S, Bisaria VS (2002) Microbial β-glucosidases: cloning, properties, and applications. Crit Rev Biotechnol 22:375–407
Beguin P, Aubert JP (1994) The biological degradation of cellulose. FEMS Microbiol Lett 13:25–58
Searchinger T, Heimlich R, Houghton RA, Dong F, Elobeid A, Fabiosa J, Tokgoz S, Hayes D, Yu TH (2008) Use of U.S. croplands for biofuels increases greenhouse gases through emissions from land-use change. Science 319:1238–1240
Chisti Y (2007) Biodiesel from microalgae. Biotechnol Adv 25:294–306
Hsieh CH, Wu WT (2009) Cultivation of microalgae for oil production with a cultivation strategy of urea limitation. Bioresource Technol 100:3921–3926
Kalinin VI, Ivanchina NV, Krasokhin VB, Makarieva TN, Stonik VA (2012) Glycosides from marine sponges (Porifera, Demospongiae): structures, taxonomical distribution, biological activities and biological roles. Mar Drugs 10:1671–1710
Fu CC, Hung TC, Chen JY, Su CH, Wu WT (2010) Hydrolysis of microalgae cell walls for production of reducing sugar and lipid extraction. Bioresource Technol 101:8750–8754
Xue DS, Chen HY, Ren YR, Yao SJ (2012) Enhancing the activity and thermostability of thermostable β-glucosidase from a marine Aspergillus niger at high salinity. Process Biochem 47:606–611
Kim YR, Kim EY, Lee JM, Kim JK, Kong IS (2013) Characterization of a novel Bacillus sp SJ-10 β-1,3-1,4-glucanase isolated from jeotgal, a traditional Korean fermented fish. Bioprocess Biosyst Eng 36:721–727
Neil JP, David EB, Emyr O, Isabel V, Jozef VB, Mahalingeshwara KB (2001) Biochemical characterization and mechanism of action of a thermostable β-glucosidase purified from Thermoascus aurantiacus. Biochem J 353:117–127
Gummadi SN (2003) What is the role of thermodynamics on protein stability? Biotechnol Bioproc E 8:9–18
Marangoni AG (2003) Enzyme kinetics: a modern approach. Wiley 146–150
Schwede T, Kopp J, Guex N, Peitsch MC (2003) SWISS-MODEL: an automated protein homology-modeling server. Nucleic Acids Res 31:3381–3385
Tina KG, Bhadra R, Srinivasan N (2007) PIC: protein interactions calculator. Nucl Acids Res 35:473–476
Gräbnitz F, Seiss M, Rücknagel KP, Staudenbauer WL (1991) Structure of the beta-glucosidase gene bglA of Clostridium thermocellum. Sequence analysis reveals a superfamily of cellulases and beta-glycosidases including human lactase/phlorizin hydrolase. Eur J Biochem 200:301–309
Sinnott ML (1990) Catalytic mechanisms of enzymic glycosyl transfer. Chem Rev 90:1171–1202
McCarter JD, Withers SG (1994) Mechanisms of enzymatic glycoside hydrolysis. Curr Opin Struc Biol 4:885–892
Kuo LC, Lee KT (2008) Cloning, expression, and characterization of two β-glucosidases from Isoflavone glycoside-hydrolyzing Bacillus subtilis natto. J Agr Food Chem 56:119–125
Paavilainen S, Hellman J, Korpela T (1993) Purification, characterization, gene cloning, and sequencing of a new β-glucosidase from Bacillus circulans subsp. Alkalophilus. Appl Environ Microb 59:927–932
González-Candelas L, Aristoy MC, Polaina J, Flors A (1989) Cloning and characterization of two genes from Bacillus polymyxa expressing β-glucosidase activity in Escherichia coli. Appl Environ Microb 55:3173–3177
Choi IS, Wi SG, Jung SR, Patel DH, Bae HJ (2009) Characterization and application of recombinant β-glucosidase (BglH) from Bacillus licheniformis KCTC 1918. J Wood Sci 55:329–334
Naz S, Ikram N, Rajoka MI, Sadaf S, Akhtar, MW (2010) Enhanced production and characterization of a β-glucosidase from Bacillus halodurans expressed in Escherichia coli. Biochemistry-Moscow 75:513–513
Hashimoto W, Miki H, Nankai H, Sato N, Kawai S, Murata K (1998) Molecular cloning of two genes for β-d-glucosidase in Bacillus sp. GL1 and identification of one as a gellan-degrading enzyme. Arch Biochem Biophys 360:1–9
Enari TM, Niku-paavola ML (1987) Enzymatic hydrolysis of cellulose: is the current theory of the mechanism of hydrolysis valid? Crit Rev Biotechnol 5:67–87
Mai Z, Yang J, Tian X, Li J, Zhang S (2013) Gene cloning and characterization of a novel salt-tolerant and glucose-enhanced β-glucosidase from a marine Streptomycete. Appl Biochem Biotech. 169:1512–1522
Zhao YH, Abraham MH (2005) Octanol/water partition of ionic species, including 544 cations. J Org Chem 70:2633–2640
Collins KD (1995) Sticky ions in biological systems. P Natl Acad Sci USA 92:5553–5557
McCarter JD, Withers SG (1994) Mechanisms of enzymatic glycoside hydrolysis. Curr Opin Struc Bio l4:885–892
Setlow B, Cabrera-Hernandez A, Cabrera-Martinez RM, Setlow P (2004) Identification of aryl-phospho-β-d-glucosidases in Bacillus subtilis. Arch Microbiol 181:60–67
Montes FJ, Battaner E, Catalan J, Galan MA (1995) Kinetics and heat-inactivation mechanism of d-amino acid oxidase. Process Biochem 30:217–224
Zale SE, Klibanov AM (1986) Why does ribonuclease irreversibly inactivate at high temperatures? Biochemistry 25:5432–5444
Vieille C, Zeikus JG (1996) Thermozymes: identifying molecular determinants of protein structural and functional stability. Trends Biotechnol 14:183–190
Daniel RM (1996) The upper limits of enzyme thermal stability. Enzyme Microb Tech 19:74–79
Matthews BW, Nicholson H, Becktel WJ (1987) Enhanced protein thermostability from site directed mutations that decrease the entropy of unfolding. P Natl Acad Sci USA 84:6663–6667
Munch O, Tritsch D (1990) Irreversible thermoinactivation of glucoamylase from Aspergillus niger and thermostabilization by chemical modification of carboxyl groups. Biochim Biophys Acta 1041:111–116
Urabe I, Nanjo H, Okada H (1973) Effect of acetylation of Bacillus subtilis α-amylase on the kinetics of heat inactivation. Biochim Biophys Acta 302:73–79
Pei XQ, Yi ZL, Tang CG, Wu ZL (2011) Three amino acid changes contribute markedly to the thermostability of β-glucosidase BglC from Thermobifida fusca. Bioresource Technol 102:3337–3342
Acknowledgments
This research was supported by iPET (Korea Institute of Planning and Evaluation for Technology in Food, Agriculture, Forestry and Fisheries), Ministry for Food, Agriculture, Forestry and Fisheries, Republic of Korea.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Lee, J.M., Kim, YR., Kim, J.K. et al. Characterization of salt-tolerant β-glucosidase with increased thermostability under high salinity conditions from Bacillus sp. SJ-10 isolated from jeotgal, a traditional Korean fermented seafood. Bioprocess Biosyst Eng 38, 1335–1346 (2015). https://doi.org/10.1007/s00449-015-1375-x
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00449-015-1375-x