The Q-loop of DrrA is involved in producing the closed conformation of the nucleotide binding domains and in transduction of conformational changes between DrrA and DrrB.

DrrA and DrrB proteins form an ATP-dependent efflux pump for doxorubicin and daunorubicin in Streptomyces peucetius. DrrA, the catalytic subunit, forms a complex with the integral membrane protein DrrB. Previous studies have provided evidence for strong interaction between these two proteins, which was found to be critical for binding of ATP to DrrA and for stability of DrrB. Chemical cross-linking experiments carried out previously showed that in the resting state of the complex DrrA and DrrB are in contact with each other. Use of a cysteine-to-amine cross-linker then allowed identification of the N-terminal cytoplasmic tail of DrrB (residues 1-53) as the primary region of contact with DrrA. In this study, single-cysteine substitutions were introduced into different domains of DrrA in a strain already containing the S23C substitution in the N-terminal tail of DrrB. By using different arm-length disulfide cross-linkers, we found that a cysteine placed in the Q-loop region of DrrA traps DrrA in the dimeric state, thus indicating that in the closed conformation the Q-loops from opposing subunits are in the proximity of each other. Furthermore, the same region of DrrA was also found to interact with the N-terminus of DrrB, although the A-A interaction was much more prominent than the A-B interaction under these conditions. On the basis of additional data shown here, we propose that the interaction of the Q-loop with the N-terminal cytoplasmic tail of DrrB identifies an important step in the communication of conformational changes between DrrA and DrrB. The significance of these findings in the mechanism of the DrrAB complex is discussed, and a model based on analyses of different conformations of DrrA and DrrB is presented.