Heterologous production and characterization of bacterial nickel/cobalt permeases

Nickel/cobalt permeases (NiCoTs, TC 2.A.52) are a rapidly growing family of structurally related membrane transporters whose members are found in Gram-negative and Gram-positive bacteria, in thermoacidophilic archaea, and in fungi. Previous studies have predicted two subclasses represented by HoxN of Ralstonia eutropha, a selective nickel transporter, and by Nh1F of Rhodococcus rhodochrous, a nickel and cobalt transporter that displays a preference for the Co ion. In the present study, NiCoT genes of five Gram-negative bacteria and one Gram-positive bacterium were cloned and heterologously expressed in Escherichia coli. Based on substrate preference in metal-accumulation assays with the recombinant strains, two of the novel NiCoTs were assigned to the Nh1F class. The remaining four NiCoTs belong to a yet unrecognized, third class. They transport both the nickel and the cobalt ion but have a significantly higher capacity for nickel. The observed substrate preferences correlate in many cases with the genomic localization of NiCoT genes adjacent to regions encoding nickel- or cobalt-dependent enzymes or enzymes involved in cobalamin biosynthesis. Alignment of 23 full-length NiCoT sequences and comparison with the available experimental data predict that substrate specificity of NiCoTs is an adaptation to specific transition metal requirements in various organisms from different taxa. (C) 2003 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.