Heterologous Expression of a Thermostable β-1,3-Galactosidase and Its Potential in Synthesis of Galactooligosaccharides
AbstractA thermostable β-1,3-galactosidase from Marinomonas sp. BSi20414 was successfully heterologously expressed in Escherichia coli BL21 (DE3), with optimum over-expression conditions as follows: the recombinant cells were induced by adding 0.1 mM of IPTG to the medium when the OD600 of the culture reached between 0.6 and 0.9, followed by 22 h incubation at 20 °C. The recombinant enzyme β-1,3-galactosidase (rMaBGA) was further purified to electrophoretic purity by immobilized metal affinity chromatography and size exclusion chromatography. The specific activity of the purified enzyme was 126.4 U mg−1 at 37 °C using ONPG (o-nitrophenyl-β-galactoside) as a substrate. The optimum temperature and pH of rMaBGA were determined as 60 °C and 6.0, respectively, resembling with its wild-type counterpart, wild type (wt)MaBGA. However, rMaBGA and wtMaBGA displayed different thermal stability and steady-state kinetics, although they share identical primary structures. It is postulated that the stability of the enzyme was altered by heterologous expression with the absence of post-translational modifications such as glycosylation, as well as the steady-state kinetics. To evaluate the potential of the enzyme in synthesis of galactooligosaccharides (GOS), the purified recombinant enzyme was employed to catalyze the transgalactosylation reaction at the lab scale. One of the transgalactosylation products was resolved as 3′-galactosyl-lactose, which had been proven to be a better bifidogenic effector than GOS with β-1,4 linkage and β-1,6 linkages. The results indicated that the recombinant enzyme would be a promising alternative for biosynthesis of GOS mainly with β-1,3 linkage. View Full-Text
Share & Cite This Article
Ding, H.; Zhou, L.; Zeng, Q.; Yu, Y.; Chen, B. Heterologous Expression of a Thermostable β-1,3-Galactosidase and Its Potential in Synthesis of Galactooligosaccharides. Mar. Drugs 2018, 16, 415.
Ding H, Zhou L, Zeng Q, Yu Y, Chen B. Heterologous Expression of a Thermostable β-1,3-Galactosidase and Its Potential in Synthesis of Galactooligosaccharides. Marine Drugs. 2018; 16(11):415.Chicago/Turabian Style
Ding, Haitao; Zhou, Lili; Zeng, Qian; Yu, Yong; Chen, Bo. 2018. "Heterologous Expression of a Thermostable β-1,3-Galactosidase and Its Potential in Synthesis of Galactooligosaccharides." Mar. Drugs 16, no. 11: 415.
Note that from the first issue of 2016, MDPI journals use article numbers instead of page numbers. See further details here.