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

A copper-transport (copYAZ) operon was cloned from the oral bacterium Streptococcus mutans JH1005. DNA sequencing showed that the operon contained three 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 with a heavy-metal-binding motif (CXCX₄CXC) at the C-terminus. CopY shared extensive homology with other bacterial negative transcriptional regulators. copA encoded a 742 aa protein that shared extensive homology with P-type ATPases. copZ encoded a 67 aa protein that also contained a heavy-metal-binding motif (CXXC) at the N-terminus. Northern blotting showed that a 3·2 kb transcript 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 inverted repeats of the promoter-operator region. Strep. mutans wild-type cells were resistant to 800 μM Cu²⁺, whereas cells of a cop knock-out mutant were killed by 200 μM Cu²⁺. Complementation of the cop knock-out mutant with the cop operon restored Cu²⁺ resistance to wild-type level. The wild-type and the mutant did not show any differences in susceptibility to other heavy metals, suggesting that the operon was specific for copper. By using a chloramphenicol acetyltransferase reporter gene fusion, the cop operon was shown to be negatively regulated by CopY and could be derepressed by Cu²⁺.