RESUMEN
CopA is a Cu(I)-exporting transmembrane P1B-type ATPase from Bacillus subtilis. It contains two N-terminal cytoplasmic domains, CopAab, which bind Cu(I) with high affinity and to form higher-order complexes with multiple Cu(I) ions. To determine the precise nature of these species, electrospray ionisation mass spectrometry (ESI-MS) under non-denaturing conditions was employed. Up to 1 Cu per CopAab resulted in Cu coordination to one or both CopAab domains. At >1 Cu/CopAab, two distinct dimeric charge state envelopes were observed, corresponding to distinct conformations, each with Cu6(CopAab)2 as its major form. The influence of the physiologically relevant low molecular weight thiol bacillithiol (BSH) on Cu(I)-binding to CopAab was assessed. Dimeric CopAab persisted in the presence of BSH, with previously undetected Cu7(CopAab)2 and Cu6(CopAab)2(BSH) forms apparent.