Attachment of Acidithiobacillus ferrooxidans onto different solid substrates and fitting through Langmuir and Freundlich equations.

Attachments of Acidithiobacillus ferrooxidans ATCC 23270 onto elemental sulfur, quartz and complex chalcopyrite were investigated by analysis of its extracellular polymeric substances as well as applying Langmuir and Freundlich equations. The two equations fitted the adsorption equilibrium data with significant correlation coefficient over 0.9. This indicated that bacterial attachment is complicated and involves Langmuir and Freundlich characterizations. Sulfur-grown cells showed the highest affinity for the three solid substrates. The investigated complex chalcopyrite possessed a higher maximum adsorption capacity for A. ferrooxidans than elemental sulfur or quartz. The Freundlich fitting parameters suggested that quartz had a weaker adsorption capacity and smaller adsorption areas than elemental sulfur or the complex chalcopyrite. It is not the content of total carbohydrates or proteins in EPS but their ratios that determine the affinity differences between cells and substrates.

Comparison of chalcopyrite bioleaching after different microbial enrichment in shake flasks.

The bioleachings of chalcopyrite ore were compared after inoculating different cultures enriched from the original acid mine drainage sample. The results showed that the higher bioleaching performance was achieved for inoculation with the enrichment D (0.5 % S, 2 % iron and 1 % chalcopyrite) compared to other enrichment systems. The generated ferric precipitation during bioleaching had a key influence on the final copper extraction. After enrichment, higher ratio of iron-oxidizer and higher ratio of sulfur-oxidizer existed in enrichment B and C, respectively. These caused the different bioleaching behaviours from other systems. Maintaining a suitable equilibrium between iron- and sulfur-oxidizers is significant to decrease ferric precipitation or postpone its formation, finally prolong efficient bioleaching period and improve copper extraction.

Metabolic changes of Acidithiobacillus caldus under Cu²(+) stress.

Metabolic changes were investigated by measuring the depletion of dissolved oxygen and the enzymatic activities of sulfur metabolism in Acidithiobacillus caldus (A. caldus) before and after copper stress. The results showed that high concentrations of Cu²(+) have an indirect negative effect on the sulfite oxidase and the APS reductase involved in sulfur metabolism when A. caldus is cultured in medium with elemental sulfur as its growth energy. This leads to a decrease in the respiration rate and the growth rate. The changes of activity are negatively correlated with the intracellular Cu²(+) concentration through an indirect interaction mechanism. A. caldus was able to induce an efflux of copper ions by forming an ATPase-dependent pump, which transported copper ions by consuming ATP. The negative effect of Cu²(+) on the bacterial metabolism could be minimized by the copper efflux when the bacteria were adapted in medium containing Cu²(+) for a long time. However, this bacterial rejuvenation became weaker when grown in medium containing higher concentrations of copper ions.

Chemical composition and antioxidant activities of Russula griseocarnosa sp. nov.

Pileus and stipe of mushroom Russula griseocarnosa from South China were analyzed separately for chemical composition and antioxidant activities. The wild mushroom species proved to have antioxidant potential, using assays of reducing power, chelating effect on ferrous ions, scavenging effect on hydroxyl free radicals, and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity. The mushroom contained very useful phytochemicals such as phenolics, flavonoids, ergosterol, and beta-carotene. The phenolic composition of R. griseocarnosa was analyzed by high-performance liquid chromatography (HPLC). The major component in R. griseocarnosa was quercetin (95.82 microg/g). The combination of bioactive substances and rich nutritional composition (high contents in protein and carbohydrates, low content in fat) in the mushroom should be useful to consumers in encouraging them to utilize the nutritive potential of this edible wild mushroom.

Bioleaching of chalcopyrite concentrate using Leptospirillum ferriphilum, Acidithiobacillus ferrooxidans and Acidithiobacillus thiooxidans in a continuous bubble column reactor.

To estimate the bioleaching performance of chalcopyrite for various hydraulic residence times (HRTs), laboratory-scale bioleaching of chalcopyrite concentrate was carried out in a continuous bubble column reactor with three different HRTs of 120, 80 and 40 h, respectively. An extraction rate and ratio of 0.578 g Cu l(-1) h(-1) and 39.7%, respectively, were achieved for an HRT of 80 h at a solids concentration of 10% (w/v). Lower bioleaching performances than this were obtained for a longer HRT of 120 h and a shorter HRT of 40 h. In addition, there was obvious competition between Leptospirillum ferriphilum and Acidithiobacillus ferrooxidans to oxidize ferrous iron, causing large compositional differences between the microbial communities obtained for the different HRTs. Leptospirillum ferriphilum and Acidithiobacillus thiooxidans were found to be the dominant microbes for the longer HRT (120 h). Acidithiobacillus ferrooxidans became the dominant species when the HRT was decreased. The proportion of Acidithiobacillus thiooxidans was comparatively constant in the microbial community throughout the three process stages.

Comparison of bioleaching behaviors of different compositional sphalerite using Leptospirillum ferriphilum, Acidithiobacillus ferrooxidans and Acidithiobacillus caldus.

Two sphalerite samples with different iron/sulphur (Fe/S) ratios, Shuikousan ore (Fe/S 0.2) and Dachang ore (Fe/S 0.52), were processed using three microbial species, Leptospirillum ferriphilum, Acidithiobacillus ferrooxidans and Acidithiobacillus caldus. Following 20 days of bioleaching in shake flask cultures, a higher zinc (Zn) extraction (96%) was achieved with Shuikousan ore than with Dachange ore (72%). The extraction efficiency increased when elemental S was added to Dachang ore to attain the same Fe/S ratio as that for Shuikousan ore. Following the addition of S, the redox potential, pH and total dissolved Fe for Dachang ore demonstrated similar behaviors to those of Shuikousan ore. Acidithiobacillus caldus and L. ferriphilum became the dominant species during the bioleaching of sphalerite with a high Fe/S ratio. In contrast, the dominant species were A. ferrooxidans and A. caldus during the bioleaching of sphalerite with a low Fe/S ratio. These results show that the Fe/S ratio has a significant influence on the bioleaching behavior of sphalerite and the composition of the microbial community.

Characterization and reconstitute of a [Fe4S4] adenosine 5'-phosphosulfate reductase from Acidithiobacillus ferrooxidans.

Adenosine 5'-phosphosulfate (APS) reductase is a key enzyme involved in the pathways of sulfate reduction and sulfide oxidation in the biological sulfur cycle. In this study, the gene of APS reductase from Acidithiobacillus ferrooxidans was cloned and expressed in Escherichia coli, the soluble protein was purified by one-step affinity chromatography to apparent homogeneity. The molecular mass of the recombinant APS reductase was determined to be 28 kDa using SDS-PAGE. According to optical and EPR spectra results of the recombinant protein confirmed that the iron-sulfur cluster inserted into the active site of the protein. Site-directed mutation for the enzyme revealed that Cys110, Cys111, Cys193, and Cys196 were in ligation with the iron-sulfur cluster. The [Fe4S4] cluster could be assembled in vitro, and exhibited electron transport and redox catalysis properties. As we know so far, this is the first report of expression in E. coli of APS reductase from A. ferrooxidans.

Expression, purification and characterization of an iron-sulfur cluster assembly protein, IscU, from Acidithiobacillus ferrooxidans.

An iron-sulfur cluster assembly protein, IscU, is encoded by the operon iscSUA in Acidithiobacillus ferrooxidans. The gene of IscU was cloned and expressed in Escherichia coli. The protein was purified by one-step affinity chromatography to homogeneity. The protein was in apo-form, the [Fe(2)S(2)] cluster could be assembled in apoIscU with Fe(2+) and sulfide in vitro, and in the presence of IscA and IscS, the IscU could utilize L: -cysteine and Fe(2+) to synthesize [Fe(2)S(2)] cluster in the protein. Site-directed mutagenesis for the protein revealed that Cys37, Asp39, Cys63 and Cys106 were involved in ligating with the [Fe(2)S(2)] cluster.

The sulfhydryl group of Cys138 of rusticyanin from Acidithiobacillus ferrooxidans is crucial for copper binding.

Rusticyanin is a small blue copper protein isolated from Acidithiobacillus ferrooxidans with extreme acid stability and redox potential. The protein is thought to be a principal component in the iron respiratory electron transport chain in this microorganism, but its exact role in electron transfer remains controversial. The gene of rusticyanin was cloned then overexpressed in Escherichia coli, the soluble protein was purified by one-step affinity chromatography to apparent homogeneity. It was reported that Cys138, His85 and His143 were important residues for copper binding, but the significance of Cys138 was not verified so far. We constructed the mutant expression plasmids of these three residues using site-directed mutagenesis. Mutant proteins were expressed in E. coli and purified with a nickel metal affinity column. The EPR and atomic absorption spectroscopy results confirmed that Cys138 was crucial for copper binding. Removal of the sulfhydryl group of Cys138 resulted in copper loss. Mutations of His85 and His143 showed little effect on copper binding.

Expression, purification and molecular modelling of the Iro protein from Acidithiobacillus ferrooxidans Fe-1.

The Iro protein was proposed to be involved in the iron respiratory electron transport chain in Acidithiobacillus ferrooxidans, it is a member of HiPIP family with the iron-sulfur cluster for electron transfer. The gene of Iro protein from A. ferrooxidans Fe-1 was cloned and then successfully expressed in Escherichia coli, finally purified by one-step affinity chromatography to homogeneity. The recombinant protein was observed to be dimer. The molecular mass of a monomer containing the [Fe4S4] cluster was 6847.35 Da by MALDI-TOF-MS. The optical and EPR spectra results of the recombinant protein confirmed that the iron-sulfur cluster was correctly inserted into the active site of the protein. Molecular modelling for the protein revealed that Cys20, Cys23, Cys32 and Cys45 were in ligation with the iron-sulfur cluster, and Tyr10 was important for the stability of the [Fe4S4] cluster. As we know, this is the first report of expression in E. coli of the Iro protein from A. ferrooxidans Fe-1.