title: Substrate binding to the multidrug transporter MepA.
abstract: MepA is a multidrug transporter from Staphylococcus aureus that confers multidrug resistance through the efflux of a wide array of hydrophobic substrates. To evaluate the ability of MepA to recognize different substrates, the dissociation constants for interactions between MepA and three of its substrates (acriflavine (Acr), rhodamine 6G (R6G), and ethidium (Et)) were measured. Given that MepA is purified in the presence of detergents and that its substrates are hydrophobic, we examined the effect of the detergent concentration on the dissociation constant. We demonstrate that all three substrates interact directly with the detergent micelles. Additionally, we find the detergent effect on the KD value to be highly substrate-dependent. The KD value for R6G is greatly influenced by the detergent, whereas the KD values for Acr and Et are only modestly affected. The effect of the inactive D183A mutant on binding was also evaluated. The D183A mutant shows lower affinity toward Acr and Et.
authors: Department of Molecular Physiology and Biological Physics, University of Virginia School of Medicine, Charlottesville, VA, 22903, United States.
authors: Christian
authors: C
authors: Banchs
authors: Department of Molecular Physiology and Biological Physics, University of Virginia School of Medicine, Charlottesville, VA, 22903, United States.
authors: Sandra
authors: S
authors: Poulos
authors: Department of Molecular Physiology and Biological Physics, University of Virginia School of Medicine, Charlottesville, VA, 22903, United States.
authors: Waroot S
authors: WS
authors: Nimjareansuk
authors: Department of Molecular Physiology and Biological Physics, University of Virginia School of Medicine, Charlottesville, VA, 22903, United States.
authors: Ye Eun
authors: YE
authors: Joo
authors: Department of Molecular Physiology and Biological Physics, University of Virginia School of Medicine, Charlottesville, VA, 22903, United States. Electronic address: sf3bb@virginia.edu.
authors: Salem
authors: S
authors: Faham
mesh_terms: D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003902:Detergents / Q000737:chemistry; D024901:Drug Resistance, Multiple, Bacterial; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D008823:Micelles; D020125:Mutation, Missense; D027701:Organic Cation Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D013211:Staphylococcus aureus / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D003902:Detergents; D005456:Fluorescent Dyes; D008823:Micelles; D027701:Organic Cation Transport Proteins
keywords: Detergent effect; Drug design; Drug resistance; Efflux; Membrane proteins