Functional analyses reveal an important role for tyrosine residues in the staphylococcal multidrug efflux protein QacA

Jingqin Wu,Karl A Hassan,Melissa H Brown,Ronald A Skurray

doi:10.1186/1471-2180-8-147

Jingqin Wu, Karl A Hassan + Show 2 more

Open Access

https://doi.org/10.1186/1471-2180-8-147

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Abstract

BackgroundThe staphylococcal QacA multidrug efflux protein confers resistance to an exceptional number of structurally unrelated antimicrobial compounds. Aromatic amino acid residues have been shown to be highly important for the transport function of several multidrug transporters and are intimately involved in multidrug binding. This study investigated the structural and functional importance of the seven tyrosine residues in QacA by examining the phenotypic effect of incorporating conservative (aromatic) and non-conservative (non-aromatic) substitutions for these residues.ResultsDetermination of the resistance profiles and analysis of drug transport assays revealed that non-conservative substitutions for most tyrosine residues influenced the QacA drug recognition spectrum. However, an aromatic residue at three tyrosine positions, 63, 410 and 429, was of importance for QacA-mediated transport and resistance to the majority of substrates tested.ConclusionA tyrosine or phenylalanine residue at amino acid positions corresponding to 63 of QacA in related drug efflux proteins is found to be highly conserved. Therefore, an aromatic side chain at this position is likely to partake in a function common to these drug transporters, such as proton translocation or essential intramolecular contacts, whereas aromatic residues at the non-conserved 410 and 429 positions are expected to mediate a QacA-specific function, possibly forming or stabilising part of the QacA drug binding region.

Highlights

The staphylococcal QacA multidrug efflux protein confers resistance to an exceptional number of structurally unrelated antimicrobial compounds
The QacA polypeptide is composed of 514 amino acid residues, organised into 14 α-helical transmembrane segments (TMS) (Figure 1) [6,7] and is classified as a member of the drug:H+ antiporter (DHA) 2 family of the major facilitator superfamily (MFS) of transport proteins [8]
Biochemical studies of TMS lining the binding region have suggested that these binding sites may be in proximity [10]

Summary

Introduction

The staphylococcal QacA multidrug efflux protein confers resistance to an exceptional number of structurally unrelated antimicrobial compounds. The QacA multidrug efflux protein confers resistance to at least 30 structurally distinct monovalent or bivalent cationic lipophilic antimicrobials from at least 12 different chemical families [1,2]. The QacA multidrug resistance protein facilitates an efflux mechanism by which strains of. The QacA polypeptide is composed of 514 amino acid residues, organised into 14 α-helical transmembrane segments (TMS) (Figure 1) [6,7] and is classified as a member of the drug:H+ antiporter (DHA) 2 family of the major facilitator superfamily (MFS) of transport proteins [8]. The drug binding region within QacA appears to contain distinct binding sites for monovalent and bivalent cationic antimicrobials [9]. The location of this residue within the binding region is flexible and appears to influence the bivalent drug recognition spectrum of the

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