Towards efficient enzymatic conversion of d-galactose to d-tagatose: purification and characterization of l-arabinose isomerase from Lactobacillus brevis
- 75 Downloads
l-arabinose isomerase (l-AI) (EC 5. 3. 1. 4. l-AI) that mediates the isomerization of d-galactose to d-tagatose was isolated from Lactobacillus brevis (MF 465792), and was further purified and characterized. Pure enzyme with molecular weight of 60.1 kDa was successfully obtained after the purification using Native-PAGE gel extraction method, which was a monomer in solution. The l-AI was found to be stable at 45–75 °C, and at pH 7.0–9.0. Its optimum temperature and pH was determined as 65 °C and 7.0, respectively. Besides, we found that Ca2+, Cu2+, and Ba2+ ions inhibited the enzyme activity, whereas the enzyme activity was significantly enhanced in the presence of Mg2+, Mn2+, or Co2+ ions. The optimum concentration of Mn2+ and Co2+ was determined to be 1 mM. Furthermore, we characterized the kinetic parameters for l-AI and determined the Km (129 mM) and the Vmax (0.045 mM min− 1) values. Notably, L. brevisl-AI exhibited a high bioconversion yield of 43% from d-galactose to d-tagatose under the optimal condition, and appeared to be a more efficient catalyst compared with other l-AIs from various organisms.
KeywordsPurification Characterization l-arabinose isomerase Lactobacillus brevis d-tagatose
Sodium dodecyl sulfate
Polyacrylamide gel electrophoresis
- L. brevis
This research was supported by the State Ethnic Affairs Commission & Ministry of Education, China. We thank Professor Shuxia Lv from Shenyang Agricultural University for helpful discussions on this manuscript.
This study was funded by the Doctoral Science Research Foundation of Liaoning Province of China (No. 20141111).
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
This article does not contain any studies with animals performed by any of the authors.
- 2.Liang M, Chen M, Liu XY, Zhai YF, Liu XW, Zhang HC, Xiao M, Wang P (2012) Bioconversion of d-galactose to d-tagatose: continuous packed bed reaction with an immobilized thermostable l-Arabinose isomerase and efficient purification by selective microbial degradation. Appl Microbiol Biotechnol 93:1469–1474CrossRefGoogle Scholar
- 9.Staudigl P, Haltrich D, Peterbauer CK (2014) l-Arabinose Isomerase and d-Xylose Isomerase from Lactobacillus reuteri: Characterization, coexpression in the food grade host Lactobacillus plantarum, and application in the conversion of d-galactose and d-glucose. J Agric Food Chem 62:1617–1624CrossRefGoogle Scholar
- 14.Suzuki S, Nira HK, Suganuma H, Suzuki C, Saito T, Yajima N (2014) Cellular fatty acid composition and exopolysaccharide contribute to bile tolerance in Lactobacillus brevis strains isolated from fermented Japanese pickles. World J Microb Biot 60:183–191Google Scholar
- 16.Xu Z, Li S, Liang JF, Feng XH, Xu H (2015) Protein purification, crystallization and preliminary X-ray diffraction analysis of l-arabinose isomerase from Lactobacillus fermentum CGMCC2921. Acta Crystallogr F71:28–33Google Scholar
- 19.Chang XY, Ying B, Zhang YL, Cao HF, Zhou T, Zhong PA, Xu B (2016) The study of food-grade induced expression and enzymatic properties of l-arabinose isomerase from Lactobacillus plantarum WU14 with high d-tagatose yield. Pol J Food Nutr Sci 7:320–337Google Scholar
- 21.Rhimi M, Bejar S (2006) Cloning, purification and biochemical characterization of metallic-ions independent and thermoactive l-arabinose isomerase from the Bacillus stearothermophilus US100 strain. BBA-Biomembr 1760:191–199Google Scholar
- 27.Kim BC, Lee YH, Lee HS, Lee DW, Choe EA, Pyun YR (2002) Cloning, expression and characterization of l-arabinose isomerasefrom Thermotoga neapolitana: bioconversion of d-galactose to d-tagatose using the enzyme. FEMS Microbiol Lett 212:121–126Google Scholar
- 31.Nakamatu T, Yamanaka K (1969) Crystallization and properties of l-arabinose isomerase from Lactobacillus gayonii. Biochim Biophys Acta 78:I56–I105Google Scholar