K-113. A Proteomic Profile of a Novel Arsenite Oxidizing Bacterium Alkalilimnicola ehrlichii strain MLHE-1T

C. Richey1, P. Chovanec1, S. Hoeft2, R. Oremland2, P. Basu1, J. Stolz1;
1Duquesne Univ., Pittsburgh, PA, 2USGS, Menlo Park, CA.

Arsenic speciation, mobility, and toxicity can be greatly influenced by microbial activity (e.g., arsenate reduction, arsenite oxidation, methylation, demethylation). The arsenite oxidizer Alkalilimnicola ehrlichii strain MLHE-1T is a haloalkaliphilic gammaproteobacterium isolated from Mono Lake, CA. It has the unique ability to grow both aerobically as a heterotroph and anaerobically as a chemolithoautotroph by coupling arsenite oxidation to nitrate reduction. The latter ability has implications for arsenic and nitrogen cycling as well as for carbon fixation in anoxic environments. The genome of MLHE-1T has been sequenced, and the operons for carbon fixation, CO oxidation, nitrate reduction, and arsenic resistance have been identified. Surprisingly, no homolog for arsenite oxidase was found, but two arsenate reductases were discovered. In addition to the dissimilatory nitrate reductase, homologs for nitric oxide reductase and nitrous oxide reductase were found. MLHE-1T lacks a homolog for nitrite reductase and nitrite is the end product of nitrate respiration. A proteomic approach was employed to understand the complex physiology of this novel bacterium. Two-dimensional PAGE gels were used to isolate proteins present in the soluble (e.g., cytosolic) and particulate (e.g., membrane) cell fractions of the bacteria grown under both oxic and anoxic conditions. Blue native gels were also used to separate individual proteins of membrane-bound complexes. Trypsin digested samples were analyzed with MALDI-TOF mass spectrometry. Proteins involved in aerobic respiration were identified in aerobically grown cells (e.g., superoxide dismutase, catalase/peroxidase I). Most of the Calvin cycle proteins (including RuBisCo) and nitrate reduction (e.g., NarH) were identified in the anaerobic fraction. Interestingly, nitrous-oxide reductase (e.g., NosZ) was a prominent protein in the anaerobic fractions. In addition, the catalytic subunit, ArrA, of one of the two putative arsenate reductases (ArrDCBAB) was found. Whether either reductase is functional has yet to be determined.