In contrast to other well-studied CBA-type heavy metal efflux systems, Cus was shown to be a tetrapartite resistance system that involves the novel periplasmic copper-binding protein CusF. These data provide additional evidence for the hypothesis that Cu(I) is directly transported from the periplasm across the outer membrane by the Cus complex.

In this article, a mechanism of arsenite [As(III)] resistance through methylation and subsequent volatization is described. Heterologous expression of arsM from Rhodopseudomonas palustris was shown to confer As(III) resistance to an arsenic-sensitive strain of Escherichia coli. ArsM catalyzes the formation of a number of methylated intermediates from As(III), with trimethylarsine as the end product. The net result is loss of arsenic, from both the medium and the cells. Because ArsM homologues are widespread in nature, this microbial-mediated transformation is proposed to have an important impact on the global arsenic cycle.As(III) ͉ ArsM ͉ methylation A s genomes are sequenced, it is becoming clear that nearly all bacteria and archaea have arsenic-resistance (ars) operons that confer resistance to arsenite [As(III)] and arsenate [As(V)] (1). The widespread occurrence of ars genes reflects the fact that arsenic is a ubiquitous environmental toxic metal. In most cases, these operons encode transport proteins that extrude As(III) from cells. In eukaryotes, As(III) detoxification involves glutathionylation coupled to removal of the As(GS) 3 complex from the cytosol by ABC transporters, such as the Saccharomyces cerevisiae Ycf1p vacuolar pump (2) or mammalian biliary extrusion pump MRP2 (3). In many mammals, including humans, an alternate metabolic fate of As(III) is methylation in the liver, followed by urinary excretion of the methylated species (4). In the past, this process was considered a detoxification mechanism (5), but more recent data suggest that the methylation actually increases toxicity by producing the more toxic monomethylarsenite [MMA(III)] and dimethylarsenite [DMA(III)], calling into question whether the process is, in fact, a detoxification process (6). An enzyme (termed Cyt19 or As3MT) that catalyzes As(III)-S-adenosylmethyltransferase activity has been identified recently in rats and humans (7-9). The enzyme has been characterized in vitro, but its physiological role is unknown.Bacteria and fungi are known to produce volatile and toxic arsines (10) but the physiological roles of arsenic methylation in microorganisms are likewise unclear, and the biochemical basis is unknown. While examining microbial genomes, we identified large number of genes for bacterial and archaeal homologues of Cyt19. We have termed a subset of these genes arsM and their protein product ArsM (As(III) S-adenosylmethyltransferase). What sets these arsM genes apart from genes for other homologues is that they are each downstream of an arsR gene, encoding the archetypal arsenic-responsive transcriptional repressor that controls expression of ars operons (11), suggesting that these ArsMs evolved to confer arsenic resistance.The gene for the 283-residue ArsM (29,656 Da) (accession no. NP948900.1) was cloned from Rhodopseudomonas palustris and expressed in an arsenic-hypersensitive strain of Escherichia coli. As(III)-resistance cells in E. coli expressing recombinant arsM correlated with conversion of medium arsenic to the methy...

The Escherichia coli zupT (formerly ygiE) gene encodes a cytoplasmic membrane protein (ZupT) related to members of the eukaryotic ZIP family of divalent metal ion transporters. Previously, ZupT was shown to be responsible for uptake of zinc. In this study, we show that ZupT is a divalent metal cation transporter of broad substrate specificity. An E. coli strain with a disruption in all known iron uptake systems could grow in the presence of chelators only if zupT was expressed. Heterologous expression of Arabidopsis thaliana ZIP1 could also alleviate iron deficiency in this E. coli strain, as could expression of indigenous mntH or feoABC. Transport studies with intact cells showed that ZupT facilitates uptake of 55 Fe 2؉ similarly to uptake of MntH or Feo. Other divalent cations were also taken up by ZupT, as shown using 57 Co 2؉ . Expression of zupT rendered E. coli cells hypersensitive to Co 2؉ and sensitive to Mn 2؉ . ZupT did not appear to be metal regulated: expression of a ⌽(zupT-lacZ) operon fusion indicated that zupT is expressed constitutively at a low level.

Transcription of the ybcZ-ylcA ylcBCD-ybdE region of the Escherichia coli K38 chromosome was analysed by Northern RNA-DNA hybridization, RT-PCR and primer extension. Transcription of a dicistronic ybcZ-ylcA mRNA and a tetracistronic ylcBCD-ybdE mRNA was induced by silver and was initiated from the sigma-70 promoters ylcAp and ylcBp. Expression of β-galactosidase activity from a Φ(ylcBp-lacZ) operon fusion was also induced by Ag M and Cu 2M , but not by Zn 2M . In-frame deletion of ybdE from the chromosome yielded a silversensitive E. coli mutant strain which did not differ in its copper resistance from its wild-type strain. On the other hand, deletion of the copA gene for the copper-exporting P-type ATPase CopA resulted in copper sensitivity, but not in silver sensitivity. A ∆ybdE ∆copA double mutant strain behaved towards copper as the ∆copA strain and towards silver as the ∆ybdE strain. Thus, in E. coli, the YlcBCD-YbdE system may be involved in silver-but not in copper resistance, and CopA may be involved in copper-but not in silver resistance.

The Escherichia coli zitB gene encodes a Zn(II) transporter belonging to the cation diffusion facilitator family. ZitB is specifically induced by zinc. ZitB expression on a plasmid rendered zntA-disrupted E. coli cells more resistant to zinc, and the cells exhibited reduced accumulation of 65 Zn, suggesting ZitB-mediated efflux of zinc.