Iron oxidase was proposed to be the initial electron acceptor from the ferrous ion in the iron oxidation of the Acidithiobacillus genus for metal bioextraction; however, its most fundamental property of direct ferrous oxidation kinetics remains undetermined due to the confusion of reaction monitor method. Here, a recombinant iron oxidase from Acidithiobacillus ferridurans Riv11 was constructed, expressed, purified, characterized, and further used to investigate the kinetics and mechanism of ferrous oxidation. This protein is more stable in an acid solution than in a neutral solution. An infrared characteristic peak around 1050 cm⁻¹ of the [Fe₄S₄] cluster was identified. The [Fe₄S₄] cluster does not affect the secondary structure of protein, but plays an important role in the stability of protein and strongly absorbs the intrinsic fluorescence of protein, resulting in a great loss of the fluorescence emission. The protein has far more absorbance than those of the iron ions in solution in the visible region; therefore, the maximum difference absorbance around 500 nm between the oxidized and reduced states of protein can be used to monitor the reaction of ferrous oxidation. Accordingly, the kinetic rate constant of the reaction was determined. Bioinformatics analysis and molecular simulation further revealed the underlying molecular mechanism that ferrous ions approach the protein at the edge of a large hydrophobic surface patch nearest to the [Fe₄S₄] cluster from a direction far from all positively charged residues, which enough enables an efficient electron transfer.

Nowadays, research on promoting the dissolution of chalcopyrite is important. As a natural symbiotic mineral of chalcopyrite, arsenopyrite will have an impact on the dissolution of chalcopyrite. This paper shows the influence of arsenopyrite on the dissolution of chalcopyrite in an acidic culture medium. The leaching results showed that adding arsenopyrite increased the leaching concentration of copper by 332 mg/L. The residues showed a decrease in sulfur through X-ray diffraction analysis (XRD) and an increase in dissolution degree through scanning electron microscope (SEM). Electrochemical experiments have shown that the rest potential of arsenopyrite is higher than that of chalcopyrite, so there is a galvanic interaction, and the impact on chalcopyrite is greater than that of arsenopyrite. The polarization curve also proves this. Under the interaction of galvanic couples, the reduction of S⁰ production and the enhancement of Cu²⁺ release can promote the dissolution of chalcopyrite. In addition, X-ray photoelectron spectrometer (XPS) analysis under the action of galvanic coupling indicates that more SO₄²⁻ is generated on the surface of chalcopyrite, replacing S_n²⁻/S⁰, and SEM shows a stronger corrosion morphology. All results confirm that the electrochemical effect between arsenopyrite and chalcopyrite promotes the dissolution of chalcopyrite in the acidic culture medium.

The genus Acidithiobacillus has been widely used in bioleaching, and novel strains in this genus, such as A. ferriphilus, have also been confirmed to possess bioleaching capabilities. In this study, an Acidithiobacillus ferriphilus strain, QBS3, was isolated from zinc-rich sulfide mine drainage using the gradient dilution method. QBS3 is a Gram-negative, 1.3 µm rod-shaped bacterium with small red colonies. It showed a high iron oxidation efficiency of 0.361 g/(L·h) and a sulfur oxidation efficiency of 0.206 g/(L·d). QBS3 has sphalerite bioleaching ability; using QBS3 for pure sphalerite bioleaching, 18.8% of zinc was extracted in 14 days at 1% pulp density. Whole genome sequencing was performed on QBS3. Functional prediction showed that 9.13% of the genes were involved in replication, recombination, and repair. Bioleaching-related genes were analyzed, including iron and sulfur oxidation genes, and carbon and nitrogen fixation genes. For iron oxidation, the Cyc2→RusA pathway and Iro→RusB pathway were found in QBS3. In terms of sulfur oxidation, QBS3 has an incomplete SOX system and lacks the SDO gene, but Rho and Trx may complement the SOX system, enabling QBS3 to oxidize sulfur. QBS3 has multiple sets of carbon fixation genes, and nitrogen fixation genes were also identified. A hypothetical sphalerite bioleaching model is proposed; this study provides a theoretical basis for the zinc sulfide ore bioleaching industry.