Cross-talk between energy supply and drug translocation in the ABC transporter Pdr5

• Applicants: Professor Dr. Lutz Schmitt; Professor Dr.-Ing. Richard Wagner
• Subject Area: Biophysics ; Biochemistry ; Structural Biology
• Term: since 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 492624709

Multidrug resistance (MDR) is a major concern in public health and in particular with the relentless rise of microbes that become resistant to multiple therapies. Among the arsenal used by microorganisms to resist to therapeutic drugs, a major threat is the use of multidrug transporters that expel many structurally unrelated drugs. Some of these polyspecific transporters use ATP as the energy source and they belong to a very large superfamily, the ATP-Binding Cassette (ABC) proteins. All members of this ABC family, primarily membrane proteins, possess conserved sequence motifs involved in ATP binding and hydrolysis and it was originally suggested that they work according to a conserved ‘unified’ mechanism of energy transduction. However, due to the versatility of function and many different topologies of ABC transporters, how and which energy is used may vary considerably, and subclasses of transporters with singular features are likely to emerge. For instance, even if all ABC transporters possess two ATP-binding sites, they are either symmetric, both sites being able to hydrolyse ATP, or asymmetric with one degenerate site, unable, or poorly able, to hydrolyse ATP. For example, Pdr5 is a full-length yeast MDR ABC transporter and the most extreme example of an ABC transporter containing a degenerated ATP binding site as all conserved motifs involved in ATP binding show exchange of the catalytical relevant amino acids. With respect to energy supply, Pdr5 seems to adapt its nucleotide dependency according to the drug transported. Moreover, recent results obtained with Pdr5 strongly support that this transporter co-transport drugs and protons and this might be relevant for other multidrug ABC transporters as well. These singularities among MDR ABC transporters warrant a detailed investigation of their molecular mechanisms with a particular focus on their energy requirement. We will therefore address central questions regarding (i) the mechanism of co-transport of protons during the transport cycle of Pdr5, (ii) specificity for certain nucleotides also in dependence of the transport substrate, (iii) continue our structural analysis of substrates and inhibitors to understand what make a substrate a substrate and an inhibitor to an inhibitor and (vi) determine the underlying dynamics and energetics during the transport cycle of Pdr5. The complementary expertise of the two teams and their knowledge of this yeast transporter as well as in electrophysiology and single molecule fluorescence techniques will be an asset to tackle these burning questions and unravel the catalytic mechanisms of Pdr5.

• DFG Programme: Research Grants