Recombinant glycerol dehydratase from Klebsiella pneumonia XJPD-Li: Induction optimization, purification and characterization

Glycerol dehydratase (GDHt) is the rate limiting enzyme in the biosynthesis of 1,3-propanediol from glycerol. The optimization of inducting process for recombinant GDHt from Klebsiella pneumoniae XJPD-Li carried out to increase specific activity and ratio of soluble form. The optimum condition was inducing under the isopropyl-β-D-thiogalactoside concentration of 0.8 mM and the temperature of 20°C for 3 h. Homogeneity of GDHt then was obtained by affinity chromatography, resulted in 2.11-fold purification and an overall yield of 47.5%. The optimum pH and reaction temperature of GDHt were pH 8.0 and 45°C, respectively. The K_m for glycerol, 1,2-propanediol, 1,2-ethanediol and coenzyme B12 were 0.48 mM, 1.43 mM, 3.07 mM, and 10.03 nM, respectively. The GDHt showed relatively stable even under temperature of 40°C and a bit blunt to oxygen. The thermo-inactivation kinetic models were fit linear under different temperatures.