Se(VI) reduction and the precipitation of Se(0) by the facultative bacterium Enterobacter cloacae SLD1a-1 are regulated by FNR

Abstract

The fate of selenium in the environment is controlled, in part, by microbial selenium oxyanion reduction and Se(0) precipitation. In this study, we identified a genetic regulator that controls selenate reductase activity in the Se-reducing bacterium Enterobacter cloacae SLD1a-1. Heterologous expression of the global anaerobic regulatory gene fnr (fumarate nitrate reduction regulator) from E. cloacae in the non-Se-reducing strain Escherichia coli S17-1 activated the ability to reduce Se(VI) and precipitate insoluble Se(0) particles. Se(VI) reduction by E. coli S17-1 containing the fnr gene occurred at rates similar to those for E. cloacae, with first-order reaction constants of k = 2.07 x 10(-2) h(-1) and k = 3.36 x 10(-2) h(-1), respectively, and produced elemental selenium particles with identical morphologies and short-range atomic orders. Mutation of the fnr gene in E. cloacae SLD1a-1 resulted in derivative strains that were deficient in selenate reductase activity and unable to precipitate elemental selenium. Complementation by the wild-type fnr sequence restored the ability of mutant strains to reduce Se(VI). Our findings suggest that Se(VI) reduction and the precipitation of Se(0) by facultative anaerobes are regulated by oxygen-sensing transcription factors and occur under suboxic conditions.

FIG. 1.

Se(VI) reduction by E. coli(pLAFR3) (□) and E. coli(pECL1e) (▪). Points and error bars represent the averages and standard deviations of triplicate experiments.

FIG. 2.

Map of the 9.76-kb HindIII fragment cloned from E. cloacae SLD1a-1. Arrows depict gene location and transcriptional direction. Lines with plasmid designation depict specific clones tested for selenate reductase activity. + or −, positive or negative for selenate reductase activity, respectively.

FIG. 3.

(A) Se(VI) reduction by E. cloacae fnr knockout mutant strain FM-1 (○) and E. cloacae SLD1a-1 wild type (•). (B) Se(VI) reduction by the SLD1a-1 wild type, the FM-1 mutant strain, and the complemented FM-1 mutant carrying pECL32. The data and error bars represent the averages and standard deviations of triplicate experiments.

FIG. 4.

SEM image of Se particles precipitated by E. cloacae (A) and E. coli(pECL1e) (B). Scale bar, 300 nm.

FIG. 5.

X-ray absorption spectra of Se particles precipitated by E. cloacae (top) and E. coli(pECL1e) (bottom). Selenium K-edge XANES spectra (A), k³-weighted χ(k) spectra (B), and the corresponding Fourier transform of the XAFS data (C) are shown.