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Biochemical characterization of recombinant benzyl alcohol dehydrogenase from Rhodococcus ruber UKMP-5M
2017
Tavakoli, Arezoo | Hamzah, Ainon
Benzyl Alcohol Dehydrogenase (BADH) is an important enzyme for hydrocarbon degradation, which can oxidize benzyl alcohols to aldehydes, while being capable of catalyzing a reversible reaction by reducing benzaldehyde. BADH is a member of medium chain alcohol dehydrogenases, in which zinc and NAD are essential for enzyme activity. This paper describes the expression, purification, and characterization of recombinant benzyl alcohol dehydrogenase, encoded by xylB gene from Rhodococcus ruber UKMP-5M. The gene has been amplified and cloned into E. coli, and the recombinant plasmid pGEMT-xylB has been digested by NdeI and HindIII to construct plasmid pET28b-xylC and then ligated into E. coli BL21 (DE3), itself induced by 0.3 mM isopropyl β-D-thiogalactoside (IPTG) at 25°C. The expressed BADH has been 38 kDa, and is purified by affinity chromatography, in which the specific activity was 30 U/mg after 17 folds purification, leading to a NAD-dependent enzyme that uses benzyl alcohol as a substrate for enzyme characterization. The final metabolite is benzaldehyde, identified by gas chromatography mass spectrometry (GC-MS). The BADH activity has been 0.7 U/mL and the optimum pH and temperature, 9.5 and 30ºC, respectively. Also the Michaelis constant (Km) and maximum velocity (Vmax) have accounted to 705 µM and 1.3 U/mL, respectively. Benzyl alcohol dehydrogenase from R. ruber UKMP-5M can be used for hydrocarbon biodegradation in contaminated sites.
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1988
Schwebach G.H. | Cafaro D. | Egan J. | Grimes M. | Michael G.
Water pollution | Nonindustrial wastewater pollution can be better managed
1991
Water pollution | State revolving funds insufficient to meet wastewater treatment needs | State revolving fund program
1992
Decontamination and energetic supply in rural installations
1992
Carbonell, V. (Escuela Tecnica Superior de Ingenieros Agronomos, Madrid (Spain). Dept. de Fisica) | Bigeriego, M. | Delgado, M.
Effects of the recirculation on the anaerobic digestion of cow manure in a biofilm reactor
1992
Sanchez Hernandez, E.P. (National Center for Scientific Research, La Habana (Cuba). Environmental Pollution Dept.) | Weiland, P. | Travieso Cordoba, L.
Common treatment of sewage sludge and animal waste slurries in covered digesters
1992
Linke, B. | Kolisch, G. (Essen Univ. (Germany). Inst. fuer Umwelttechnik)
Controlling volatile organic compound emissions from industrial wastewater
1990
Elliott, Jeffrey | Watkins, Sheryl
Treatment of microbial contaminants in potable water supplies
1989
Troyan, Jerrold J. | Hansen, Sigurd P.
Composting municipal sludge
1988
Benedict, Arthur H. | Epstein, Eliot | Alpert, Joel E.
