Papers by Keyword: Rusticyanin

Abstract: Recombinant rusticyanin was produced in Pichia pastoris, then purified and immobilized on Sepharose CL-4B with periodate activation. Cellular lysate of acidophilic Acidithiobacillus ferrooxidans was applied to an affinity column with immobilized rusticyanin. Rusticyanin-binding proteins, separated using 1D PAGE and identified by mass spectrometry, included anticipated interacting partners, such as cytochromes Cyc1 and Cyc2, which are involved in the downhill electron pathway from ferrous iron to oxygen. However, the results indicate that rusticyanin’s functional protein-protein interaction (PPI) network could be more complex than expected, including various proteins involved in different cellular processes. Although affinity purification coupled to mass spectrometry should mostly detect proteins that bind stably, and thus are likely participants in functional in vivo PPIs, further verification is needed to exclude non-functional interactants. Nevertheless, our preliminary PPI data confirm some previous experimental findings and indicate potentially fruitful directions for probing additional roles of rusticyanin in sulfur metabolism, copper resistance, anaerobic iron reduction, iron transport, and oxidative stress in extreme acidophiles.

Abstract: It has long been recognized that isolates of iron- and sulfur-oxidizing acidophiles referred to as “Acidithiobacillus ferrooxidans” probably include more than one species, on the basis of differences in chromosomal GC contents and 16S rRNA gene sequences. Phylogenetic heterogeneity among these isolates was confirmed by phylogenetic analysis using the sequences of the 16S-23S intergenic spacers (ITS). Two main groups have been identified: the first includes the type strain of Acidithiobacillus ferrooxidans and a second comprises a cluster of newly isolated strains that have 98.5% 16S rRNA gene sequence identity with the type strain. Given that the new group of isolates have GC contents of 56 mol% as opposed to 58.8 mol% for At. ferrooxidansT, and that they share only 37% homologous DNA, these were given the new species name Acidithiobacillus ferrivorans. Further studies showed that, while strains of At. ferrivorans have many physiological traits in common with At. ferrooxidans, they also differ in some key characteristics. These include the ability to grow at temperatures as low as 4°C (as opposed to the lower limit of between 10 and 12°C for At. ferrooxidans) and the greater sensitivity of At. ferrivorans to low pH (minimum of 1.9 for growth as opposed to 1.3 for At. ferrooxidansT). Important genotypic differences include the fact that all strains of At. ferrivorans do not contain the archetypal rusticyanin gene (rusA), rather most contain a rusA homologue (rusB). Furthermore, the high potential iron-sulfur protein-encoding gene of all At. ferrivorans strains analyzed is more similar to the iro than to the hip gene characterized in At. ferrooxidansT. These results suggest that the iron oxidation pathways are different in At. ferrivorans and At. ferrooxidans.

Abstract: Growth on ferrous iron of a new isolate of the halotolerant acidophile “Thiobacillus prosperus” occurred with a substrate oxidation rate similar to that of Acidithiobacillus ferrooxidans, but with a requirement for salt (NaCl). These observations contrast with the previous description of “T. prosperus” in which a salt requirement was not noted and growth on ferrous iron was described as poor. As well as similar capacities for iron oxidation, these species were shown to possess similar clusters of genes (the rus operon) that encode proteins likely to be involved in transfer of electrons from ferrous iron. There were some differences in the organization of the genes and one of them that encodes a cytochrome c in At. ferrooxidans was absent from the “T. prosperus” cluster.