American Chemical Society
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The Signaling Interface of the Yeast Multidrug Transporter Pdr5 Adopts a Cis Conformation, and There Are Functional Overlap and Equivalence of the Deviant and Canonical Q-Loop Residues

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journal contribution
posted on 2010-06-01, 00:00 authored by Neeti Ananthaswamy, Robert Rutledge, Zuben E. Sauna, Suresh V. Ambudkar, Elliot Dine, Emily Nelson, Di Xia, John Golin
ABC transporters are polytopic proteins. ATP hydrolysis and substrate transport take place in separate domains, and these activities must be coordinated through a signal interface. We previously characterized a mutation (S558Y) in the yeast multidrug transporter Pdr5 that uncouples ATP hydrolysis and drug transport. To characterize the transmission interface, we used a genetic screen to isolate second-site mutations of S558Y that restore drug transport. We recovered suppressors that restore drug resistance; their locations provide functional evidence for an interface in the cis rather than the trans configuration indicated by structural and cross-linking studies of bacterial and eukaryotic efflux transporters. One mutation, E244G, defines the Q-loop of the deviant portion of NBD1, which is the hallmark of this group of fungal transporters. When moved to an otherwise wild-type background, this mutation and its counterpart in the canonical ATP-binding site Q951G show a similar reduction in drug resistance and in the very high basal-level ATP hydrolysis characteristic of Pdr5. A double E244G, Q951G mutant is considerably more drug sensitive than either of the single mutations. Surprisingly, then, the deviant and canonical Q-loop residues are functionally overlapping and equivalent in a strikingly asymmetric ABC transporter.