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, cutC, cutD, cutE, and cutF. A mutation in one or more of these genes results 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 ion Homeostasis: Molecular Biology and Chemistry, 1989). Copper-sensitive cutC and cutF mutants were transformed with a genomic library of E. coli, and copper-tolerant transformants were selected. Two distinct clones were identified, each of which partially restores copper tolerance in both the cutC and cutF mutants of E. coli. Subcloning, physical mapping, and sequence analysis have revealed that the cutC gene is located at 42.15 min on the E. coli genome and encodes a cytoplasmic protein 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 identified by Silhavy and coworkers CW. B, Snyder, L. J. B. Davis, P. N. Danese, C. L. Cosma, and T. J. Silhavy, J. Bacteriol. 177:4216-4223, 1995). Results from the genetic mapping of the copper-sensitive mutations in the cutF mutant and sequencing of the cutC and cutF (nlpE) alleles 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 mutations in both the genes appear to be required for the copper-sensitive phenotype in each mutant.