title: Detection of protein-protein interactions in the alkanesulfonate monooxygenase system from Escherichia coli.
abstract: The two-component alkanesulfonate monooxygenase system utilizes reduced flavin as a substrate to catalyze a unique desulfonation reaction during times of sulfur starvation. The importance of protein-protein interactions in the mechanism of flavin transfer was analyzed in these studies. The results from affinity chromatography and cross-linking experiments support the formation of a stable complex between the flavin mononucleotide (FMN) reductase (SsuE) and monooxygenase (SsuD). Interactions between the two proteins do not lead to overall conformational changes in protein structure, as indicated by the results from circular dichroism spectroscopy in the far-UV region. However, subtle changes in the flavin environment of FMN-bound SsuE that occur in the presence of SsuD were identified by circular dichroism spectroscopy in the visible region. These data are supported by the results from fluorescent spectroscopy experiments, where a dissociation constant of 0.0022 +/- 0.0010 muM was obtained for the binding of SsuE to SsuD. Based on these studies, the stoichiometry for protein-protein interactions is proposed to involve a 1:1 monomeric association of SsuE with SsuD.
authors.affiliation: Department of Chemistry and Biochemistry, Auburn University, 179 Chemistry Building, Auburn, AL 36849. ellishr@auburn.edu.
authors.forename: Kholis
authors.initials: K
authors.lastname: Abdurachim
authors.forename: Holly R
authors.initials: HR
authors.lastname: Ellis
mesh_terms: D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism*; D038181:FMN Reductase / Q000737:chemistry / Q000378:metabolism*; D005415:Flavins / Q000378:metabolism; D006899:Mixed Function Oxygenases; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding
chemical_list: D029968:Escherichia coli Proteins; D005415:Flavins; D006899:Mixed Function Oxygenases; C499579:SsuD protein, E coli; D038181:FMN Reductase; C447459:SsuE protein, E coli; D009247:NADH, NADPH Oxidoreductases