Blue-Copper Proteins: Expression of Coding Genes from Sulfobacillus Spp. and Iron Oxidation in Column Bioleaching Tests

摘要

In bioleaching, the chemiolithotrophic community plays an important role as oxidizers of sulfur compounds and ferrous iron. Ferrous iron oxiding microorganisms are key players in the process, as ferric iron is absolutely required to solubilize metal sulfide ores. Members of the Sulfobacillus genus (able to oxidize ferrous iron) were predominant (22 - 95%) in a chalcopyrite bioleaching columns test. In order to obtain new insight about the mechanism of iron oxidation in Sulfobacillus we investigated the presence and expression of genes potentially related to iron oxidation by Sulfobacillus, especially the group of the so-called blue-copper proteins rusticyanin (rus) and sulfocyanin (soxE) in the course of the experiment. The physicochemical parameters and the population dynamics were monitored periodically in the columns and the metatranscriptome was analyzed by using pyro-sequencing. The average temperature inside the column ranged from 22 to 57°C and the Fe(II) oxidation rate at 45°C varied between 8 and 42 mg L~(-1)h~(-1) along 300 days of operation. The metatranscriptomic analysis reveals an over-expression of 9-13 folds of the putative rus and soxE genes in four strains of Sulfobacillus spp. when the Sulfobacillus proportion in the column was > 80% and the Fe(II) oxidation rate measured at 45°C reached 10 mg L~(-1)h~(-1). Some cytochromes from the electron transport chain were also over-expressed, on a range of 7 - 10 folds under those operational conditions. These results support the hypothetical participation of bluecopper proteins in the iron oxidation pathway of Sulfobacilli. Culture assays and more specific expression analysis are necessary in order to confirm this hypothesis. In addition, we attempt to establish the relationship between rusticyanin and sulfocyanin genes and perform a protein sequence analysis that allows us to infer the actual function of these proteins in Sulfobacillus species.