Abstract
Resistance to arsenite (As(III)) by cells is generally accomplished by arsenite efflux permeases from Acr3 or ArsB unrelated families. We analyzed the function of three Acr3 proteins from Corynebacterium glutamicum, CgAcr3-1, CgAcr3-2, and CgAcr3-3. CgAcr3-1 conferred the highest level of As(III) resistance and accumulation in vivo. CgAcr3-1 was also the most active when everted membranes vesicles from Escherichia coli or C. glutamicum mutants were assayed for efflux with different energy sources. As(III) and antimonite (Sb(III)) resistance and accumulation studies using E. coli or C. glutamicum arsenite permease mutants clearly show that CgAcr3-1 is specific for As(III). In everted membrane vesicles expressing CgAcr3-1, dissipation of either the membrane potential or the pH gradient of the proton motive force did not prevent As(III) uptake, whereas dissipation of both components eliminated uptake. Further, a mutagenesis study of CgAcr3-1 suggested that a conserved cysteine and glutamate are involved in active transport. Therefore, we propose that CgAcr3-1 is an antiporter that catalyzes arsenite-proton exchange with residues Cys129 and Glu305 involved in efflux.
Highlights
CgAcr3-1 is an arsenite permease that catalyzes As(III) efflux from Corynebacterium glutamicum
The observed As(III) resistance levels obtained for the mutants indicate that Cgacr3-1 and Cgacr3-2 are involved in arsenite resistance in C. glutamicum, whereas Cgacr3-3 is not (Fig. 1A)
We used C. glutamicum 2⌬ars as the recipient strain for homologous complementation analysis of the CgAcr3 proteins, whereas E. coli AW3110 was used for CgAcr3 heterologous complementation analysis
Summary
CgAcr is an arsenite permease that catalyzes As(III) efflux from Corynebacterium glutamicum. As(III) and antimonite (Sb(III)) resistance and accumulation studies using E. coli or C. glutamicum arsenite permease mutants clearly show that CgAcr is specific for As(III). The properties of a more distant Acr homologue from Shewanella oneidensis was examined recently [14]; this protein confers “in vivo” resistance to As(V) but not As(III), and the purified protein binds only As(V), indicating that this protein is not a true Acr orthologue Fungal members of this family include the ScAcr3p metalloid efflux protein from Saccharomyces cerevisiae, which was proposed to be selective for As(III) over Sb(III) [15]. While this study was in progress, a recent report showed Sb(III) accumulation by yeast everted vesicles expressing Acr protein [17]. Taking into account the results, we propose a model for the permease that is consistent with As(III) efflux data
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