期刊
JOURNAL OF BIOLOGICAL CHEMISTRY
卷 294, 期 13, 页码 4934-4945出版社
ELSEVIER
DOI: 10.1074/jbc.RA118.006316
关键词
copper; Pseudomonas; homeostasis; metal ion-protein interaction; metalloprotein; metal sensor; metallochaperone; stress response; transcriptional regulation
资金
- National Institutes of Health [R01GM114949]
Copper homeostasis in pathogenic bacteria is critical for cuproprotein assembly and virulence. However, in vivo biochemical analyses of these processes are challenging, which has prevented defining and quantifying the homeostatic interplay between Cu+-sensing transcriptional regulators, chaperones, and sequestering molecules. The cytoplasm of Pseudomonas aeruginosa contains a Cu+-sensing transcriptional regulator, CueR, and two homologous metal chaperones, CopZ1 and CopZ2, forming a unique system for studying Cu+ homeostasis. We found here that both chaperones exchange Cu+, albeit at a slow rate, reaching equilibrium after 3 h, a time much longer than P. aeruginosa duplication time. Therefore, they appeared as two separate cellular Cu+ pools. Although both chaperones transferred Cu+ to CueR in vitro, experiments in vivo indicated that CopZ1 metallates CueR, eliciting the translation of Cu+ efflux transporters involved in metal tolerance. Although this observation was consistent with the relative Cu+ affinities of the three proteins (CopZ1 < CueR < CopZ2), in vitro and in silico analyses also indicated a stronger interaction between CopZ1 and CueR that was independent of Cu+. In contrast, CopZ2 function was defined by its distinctly high abundance during Cu2+ stress. Under resting conditions, CopZ2 remained largely in its apo form. Metal stress quickly induced CopZ2 expression, and its holo form predominated, reaching levels commensurate with the cytoplasmic Cu+ levels. In summary, these results show that CopZ1 acts as chaperone delivering Cu+ to the CueR sensor, whereas CopZ2 functions as a fast-response Cu+-sequestering storage protein. We propose that equivalent proteins likely play similar roles in most bacterial systems.
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