Characterization of a copper-transport operon, copYAZ, from Streptococcus mutans

A copper-transport (copYAZ) operon was cloned from the oral bacterium Strep tococcus mutans JH1005. DNA sequencing showed that the operon contained thr ee genes (copY, copA and copZ), which were flanked by a single promoter and a factor-independent terminator. copy encoded a small protein of 147 aa wi th a heavy-metal-binding motif (CXCX4CXC) at the C-terminus. CopY Shared ex tensive homology with other bacterial negative transcriptional regulators. copA encoded a 742 aa protein that shared extensive homology with P-type AT Pases. copZ encoded a 67 aa protein that also contained a heavy-metal-bindi ng motif (CXXC) at the N-terminus. Northern blotting showed that a 3.2 kb t ranscript was produced by Cu2+-induced Strep. mutans cells, suggesting that the genes were synthesized as a polycistronic message. The transcriptional start site of the cop operon was mapped and shown to lie within the invert ed repeats of the promoter-operator region. Strap. mutans wild-type cells w ere resistant to 800 muM Cu2+, whereas cells of a cop knock-out mutant were killed by 200 muM Cu2+. Complementation of the cop knock-out mutant with t he cop operon restored Cu2+ resistance to wild-type level. The wild-type an d the mutant did not show any differences in susceptibility to other heavy metals, suggesting that the operon was specific for copper. By using a chlo ramphenicol acetyltransferase reporter gene fusion, the cop operon was show n to be negatively regulated by CopY and could be derepressed by Cu2+.