J. Bacteriol., 10 1996, 5699-5705, Vol 178, No. 19
Copyright © 1996, American Society for Microbiology

Gene overexpression, purification, and identification of a desulfurization enzyme from Rhodococcus sp. strain IGTS8 as a sulfide/sulfoxide monooxygenase

B Lei and SC Tu
Department of Biochemical and Biophysical Sciences, University of Houston, Texas 77204-5934, USA.

The oxidation of dibenzothiophene to dibenzothiophene sulfone has been linked to the enzyme encoded by the sox/dszC gene from Rhodococcus sp. strain IGTS8 (S. A. Denome, C. Oldfield, L. J. Nash, and K. D. Young, J. Bacteriol. 176:6707-6717, 1994; C. S. Piddington, B. R. Kovacevich, and J. Rambosek, Appl. Environ. Microbiol. 61:468-475, 1995). However, this enzyme has not been characterized, and the type of its catalytic activity remains unclassified. In this work, the sox/dszC gene was overexpressed in Escherichia coli, a procedure for the purification of the expressed enzyme was developed, and the properties of and the reactions catalyzed by the purified enzyme were characterized. This enzyme binds one flavin mononucleotide (Kd, 7 micrometers) or reduced flavin mononucleotide (FMNH2) (Kd < 10(-8) M) per 90,200-Da homodimer, and FMNH2 is an essential cosubstrate for its activity. Patterns of product formation were examined under different FMNH2 availabilities, and results indicate that this enzyme catalyzes a stepwise conversion of dibenzothiophene to the corresponding sulfoxide and subsequently to the sulfone. On the basis of isotope labeling patterns with H2(18)O and 18O2, dibenzothiophene sulfoxide and sulfone obtained their oxygen atom(s) from molecular oxygen rather than water in their formation from dibenzothiophene. The enzyme also utilizes benzyl sulfide and benzyl sulfoxide as substrates. Hence, it is identified as a sulfide/sulfoxide monooxygenase. This monooxygenase is similar to the microsomal flavin- containing monooxygenase but is unique among microbial flavomonooxygenases in its ability to catalyze two consecutive monooxygenation reactions.

This article has been cited by other articles:

• Sucharitakul, J., Chaiyen, P., Entsch, B., Ballou, D. P. (2006). Kinetic Mechanisms of the Oxygenase from a Two-component Enzyme, p-Hydroxyphenylacetate 3-Hydroxylase from Acinetobacter baumannii. J. Biol. Chem. 281: 17044-17053 [Abstract] [Full Text]  
• Erwin, K. N., Nakano, S., Zuber, P. (2005). Sulfate-Dependent Repression of Genes That Function in Organosulfur Metabolism in Bacillus subtilis Requires Spx. J. Bacteriol. 187: 4042-4049 [Abstract] [Full Text]  
• Li, F., Xu, P., Feng, J., Meng, L., Zheng, Y., Luo, L., Ma, C. (2005). Microbial Desulfurization of Gasoline in a Mycobacterium goodii X7B Immobilized-Cell System. Appl. Environ. Microbiol. 71: 276-281 [Abstract] [Full Text]  
• Van Hamme, J. D., Fedorak, P. M., Foght, J. M., Gray, M. R., Dettman, H. D. (2004). Use of a Novel Fluorinated Organosulfur Compound To Isolate Bacteria Capable of Carbon-Sulfur Bond Cleavage. Appl. Environ. Microbiol. 70: 1487-1493 [Abstract] [Full Text]  
• Sutherland, T. D., Horne, I., Russell, R. J., Oakeshott, J. G. (2002). Gene Cloning and Molecular Characterization of a Two-Enzyme System Catalyzing the Oxidative Detoxification of {beta}-Endosulfan. Appl. Environ. Microbiol. 68: 6237-6245 [Abstract] [Full Text]  
• Woodmansee, A. N., Imlay, J. A. (2002). Reduced Flavins Promote Oxidative DNA Damage in Non-respiring Escherichia coli by Delivering Electrons to Intracellular Free Iron. J. Biol. Chem. 277: 34055-34066 [Abstract] [Full Text]  
• Duarte, G. F., Rosado, A. S., Seldin, L., de Araujo, W., van Elsas, J. D. (2001). Analysis of Bacterial Community Structure in Sulfurous-Oil-Containing Soils and Detection of Species Carrying Dibenzothiophene Desulfurization (dsz) Genes. Appl. Environ. Microbiol. 67: 1052-1062 [Abstract] [Full Text]  
• Matsubara, T., Ohshiro, T., Nishina, Y., Izumi, Y. (2001). Purification, Characterization, and Overexpression of Flavin Reductase Involved in Dibenzothiophene Desulfurization by Rhodococcus erythropolis D-1. Appl. Environ. Microbiol. 67: 1179-1184 [Abstract] [Full Text]  
• Bohuslavek, J., Payne, J. W., Liu, Y., Bolton, H. Jr., Xun, L. (2001). Cloning, Sequencing, and Characterization of a Gene Cluster Involved in EDTA Degradation from the Bacterium BNC1. Appl. Environ. Microbiol. 67: 688-695 [Abstract] [Full Text]  
• Xun, L., Sandvik, E. R. (2000). Characterization of 4-Hydroxyphenylacetate 3-Hydroxylase (HpaB) of Escherichia coli as a Reduced Flavin Adenine Dinucleotide-Utilizing Monooxygenase. Appl. Environ. Microbiol. 66: 481-486 [Abstract] [Full Text]  
• Kertesz, M. A., Schmidt-Larbig, K., Wüest, T. (1999). A Novel Reduced Flavin Mononucleotide-Dependent Methanesulfonate Sulfonatase Encoded by the Sulfur-Regulated msu Operon of Pseudomonas aeruginosa. J. Bacteriol. 181: 1464-1473 [Abstract] [Full Text]  
• Payne, J. W., Bolton, H. Jr., Campbell, J. A., Xun, L. (1998). Purification and Characterization of EDTA Monooxygenase from the EDTA-Degrading Bacterium BNC1. J. Bacteriol. 180: 3823-3827 [Abstract] [Full Text]