Crystallization and preliminary X-ray crystallographic studies of the alkanesulfonate FMN reductase from Escherichia coli
- Department of Chemistry and Biochemistry, Auburn University, Auburn, AL 36849 (United States)
- Center for Structural Biology, Wake Forest University School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157 (United States)
Crystallization of the native and SeMet FMN reductase protein of the E. coli alkanesulfonate monooxygenase two-component enzyme system is reported. The alkanesulfonate FMN reductase (SsuE) from Escherichia coli catalyzes the reduction of FMN by NADPH to provide reduced flavin for the monooxygenase (SsuD) enzyme. The vapor-diffusion technique yielded single crystals that grow as hexagonal rods and diffract to 2.9 Å resolution using synchrotron X-ray radiation. The protein crystallizes in the primitive hexagonal space group P622. The SsuE protein lacks any cysteine or methionine residues owing to the role of the SsuE enzyme in the acquisition of sulfur during sulfate starvation. Therefore, substitution of two leucine residues (Leu114 and Leu165) to methionine was performed to obtain selenomethionine-containing SsuE for MAD phasing. The selenomethionine derivative of SsuE has been expressed and purified and crystals of the protein have been obtained with and without bound FMN. These preliminary studies should lead to the structure solution of SsuE. It is anticipated that this new protein structure will provide detailed structural information on specific active-site regions of the protein and insight into the mechanism of flavin reduction and transfer of reduced flavin.
- OSTI ID:
- 22356159
- Journal Information:
- Acta Crystallographica. Section F, Vol. 61, Issue Pt 9; Other Information: PMCID: PMC1978109; PMID: 16511173; PUBLISHER-ID: pu5096; OAI: oai:pubmedcentral.nih.gov:1978109; Copyright (c) International Union of Crystallography 2005; Country of input: International Atomic Energy Agency (IAEA); ISSN 1744-3091
- Country of Publication:
- United Kingdom
- Language:
- English
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