ABSTRACT
Background: Benzaldehyde dehydrogenase [BZDH] is encoded by the xylC that catalyzes the conversion of benzaldehyde into benzoate in many pathways such as toluene degradation
Objectives: In this study, the xylC gene from Rhodococcus ruber UKMP-5M was expressed in Escherichia coli, purified, and characterized
Materials and Methods: The xylC was amplified and cloned in E. coli. The recombinant plasmid pGEMT-xylC was digested by NdeI and HindIII to construct plasmid pET28b-xylC and transformed in E. coli BL21 [DE3]. Expression of the recombinant protein was induced by 1 mM isopropyl beta-D-thiogalactoside [IPTG] at 37[degree]C. The BZDH was purified by ion exchange chromatography, in which the product was an NAD-dependent enzyme using benzaldehyde as a substrate for enzyme characterization. The end metabolite was identified via gas chromatography mass spectrometry [GC-MS]
Results: The recombinant BZDH is 27 kDa, purified by ion exchange chromatography. The activity of BZDH was 9.4 U.microL[-1] The optimum pH and temperature were 8.5 and 25[degree]C, respectively. The Michaelis constant [K[m]] and maximum velocity [V[max]] were 4.2 mM and 19.7 U.mL[-1], respectively. The metabolite of BZDH was benzene carboxylic acid as determined by GC-MS analysis
Conclusions: BZDH has the ability to degrade benzaldehyde to less toxic compounds. The BZDH is a critical enzyme for the degradation of aromatic hydrocarbons in Rhodococcus sp. The BZDH from R. ruber UKMP-5M is showed similar function with other aldehyde dehydrogenases