Background: Fungal multidrug efflux pumps possess a degenerate nucleotide-binding site (NBS). Results: Restoring all the nonconserved amino acids in the degenerate NBS of Pdr5 leads to complete loss of function of the protein. Conclusion:The degenerate NBS is essential and acts as a structural platform supporting the canonical NBS. Significance: This is the first study dealing with the entire degenerate NBS and its functional role.

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A subset of the family of ATP-binding cassette (ABC) transporters has been in focus owing to their involvement in conferring multidrug resistance in cancer cells and among immune compromised individuals. Saccharomyces cerevisiae is protected against xenobiotics by similar machineries that are part of the pleitropic drug resistance (PDR) network. The ABC transporter Pdr5 is an important member of this PDR network in yeast and is involved in cellular detoxification by the efflux of a wide variety of drugs and substrates. In this review, we focus on the aspects of detergent effects and the degeneracy in conserved sequences that is observed in the nucleotide binding domains of Pdr5 and discuss their functional relevance.

The pleiotropic drug resistance network in budding yeast presents a first line of defense against xenobiotics, which is formed by primary and secondary active membrane transporters. Among these transporters, the ABC transporter Pdr5 is a key component, because it confers resistance against a broad spectrum of such cytotoxic agents. Furthermore, it represents a model system for homologous transporters from pathogenic fungi and has been intensively studied in the past. In addition to other mutational studies, the S1360F mutation of Pdr5 was found to modulate substrate specificity and resistance. Notably, in the S1360F background, the resistance against the immunosuppressant FK506 is drastically increased. We present a detailed analysis of this mutation that is located in the predicted cytosolic part of transmembrane helix 11. Our data demonstrate that kinetic and thermodynamic parameters of the S1360F mutant are similar to those of the wild-type protein, except for FK506-inhibited ATPase activity and the degree of competitive inhibition. In summary, our results indicate that the S1360F mutation within the transmembrane domain interferes drastically with the ability of the nucleotide-binding domains to hydrolyze ATP by interfering with interdomain crosstalk.