IDENTIFICATION OF CUTC AND CUTF (NLPE) GENES INVOLVED IN COPPER TOLERANCE IN ESCHERICHIA-COLI

It has been suggested previously that copper transport in Escherichia coli is mediated by the products of at least six genes, cutA, cutB, cu tC, cutD, cutE, and cutF. A mutation in one or more of these genes res ults in an increased copper sensitivity (D. Rouch, J. Camakaris, and B . T. O. Lee, p. 469-477, in D. H. Hamer and D. R. Winge, ed., Metal io n Homeostasis: Molecular Biology and Chemistry, 1989). Copper-sensitiv e cutC and cutF mutants were transformed with a genomic library of E. coli, and copper-tolerant transformants were selected. Two distinct cl ones were identified, each of which partially restores copper toleranc e in both the cutC and cutF mutants of E. coli. Subcloning, physical m apping, and sequence analysis have revealed that the cutC gene is loca ted at 42.15 min on the E. coli genome and encodes a cytoplasmic prote in of 146 amino acids and that the cutF gene is located at 4.77 min on the E. coli genome and is allelic to the nlpE gene independently iden tified by Silhavy and coworkers CW. B, Snyder, L. J. B. Davis, P. N. D anese, C. L. Cosma, and T. J. Silhavy, J. Bacteriol. 177:4216-4223, 19 95). Results from the genetic mapping of the copper-sensitive mutation s in the cutF mutant and sequencing of the cutC and cutF (nlpE) allele s from both cutC and cutF mutants indicate that both the cutC and cutF mutants are in fact double mutants altered in these two genes, and mu tations in both the genes appear to be required for the copper-sensiti ve phenotype in each mutant.