(157bi) Engineering the Bioavailability of Sulfur to Control Iron Redox States in Acidithiobacillus Ferrooxidans

The biomining microorganism, Acidithiobacillus ferrooxidans, is an acidophilic chemolithoautotroph with three major metabolic pathways to utilize iron, sulfur, and hydrogen as electron donors. Biotechnological application of this bacterium, as well as other bacteria found with it in natural consortia, is important for the global industrial biomining of approximately 5% of gold and 15% of copper¹. The iron and sulfur metabolisms have been well-studied as the interaction of A. ferrooxidans with metal sulfides and other sources of metal, such as e-waste, leads to the dissolution of metal cations. However, as most studies with this organism investigate growth with a single energy source, we have exploited the ability of A. ferrooxidans to oxidize iron, sulfur, and pyrite simultaneously to enhance the corrosion of stainless steels. This systems-level approach has allowed us to demonstrate that the optimization of substrate oxidation via the manipulation of cultivation conditions combined with genetic engineering can be used to for the microbially influenced corrosion of complex metal containing matrices².

Despite the favorable bioenergetics associated with sulfur oxidation compared to ferrous iron, the oxidation of sulfur involves an extended lag phase where A. ferrooxidans produces the necessary extracellular polymeric substances and proteins needed to attach to the hydrophobic surface of the sulfur particle. As a result of this lag phase, when both ferrous iron and sulfur are present in solution, iron oxidation behavior precedes sulfur oxidation for iron-adapted A. ferrooxidans cells, leading to the long-held conclusion that A. ferrooxidans is an iron-preferred species. We discovered a class of biocompatible surfactants, lignosulfonates, that could be used to formulate sulfur to have more hydrophilic properties and disperse into aqueous solution.

With this novel dispersed sulfur formulation, we achieved rapid utilization of sulfur indicating that there was no substrate preference for iron in A. ferrooxidans, if the provided sulfur was already bioavailable³. The observed iron oxidation behavior in the bacteria was concentration dependent on dispersed sulfur. At higher concentrations of dispersed sulfur, the iron oxidation rates were found to be slower until at concentrations above 5 g/L, ferrous iron oxidation did not reach completion and was replaced with ferric iron reduction. We investigated this iron reduction behavior further and found that aerobic reduction of ferric iron was not dependent on pH if the metabolic state of A. ferrooxidans was such that the downhill iron oxidation pathway was inactive. Therefore, by manipulating the bioavailability of sulfur, the interplay between the iron and sulfur metabolisms within A. ferrooxidans enable the tunable control of iron redox states. This new level of control can be used to manipulate the performance of the cells for future biotechnology applications.

1 Banerjee, I., Burrell, B., Reed, C., West, A. C. & Banta, S. Metals and minerals as a biotechnology feedstock: engineering biomining microbiology for bioenergy applications. Curr Opin Biotechnol 45, 144-155, doi:10.1016/j.copbio.2017.03.009 (2017).

2 Inaba, Y., West, A. C. & Banta, S. Enhanced microbial corrosion by Acidithiobacillus ferrooxidans through the manipulation of substrate oxidation and genetic engineering. (Submitted).

3 Inaba, Y., West, A. C. & Banta, S. Bioavailability of elemental sulfur drives substrate utilization behavior in Acidithiobacillus ferrooxidans. (In preparation).