Interactions of Microbial Siderophores with Titanic Ions and Titanium-Bearing Minerals

Abstract

Transition metals play an important role in many biological processes. Iron is essential for almost every organism, but its bioavailability is limited due to the low solubility of Fe(III) in aqueous environments. Microbial siderophores help solubilize and sequester iron(III). In solution, siderophores like desferrioxamine B (DFOB) are also avid binders of Ti(IV). Ti(IV) is chemically similar to Fe(III), and the use of usually-inert TiO2 is increasing in products such as sunscreens and paint. The surface of titanium metal in joint replacements and implants is oxidized to form TiO2. Microbial siderophores bind to normally inert TiO2 and this binding can affect the solubility of Ti(IV). Dissolution might render Ti(IV) biologically available, and might interfere with Fe(III) biogeochemical cycling, as well as impact biofouling in marine, medicinal, and industrial applications. This research explores how siderophores interact with Ti(IV) in aqueous solutions and can solubilize Ti(IV) from the surface of solid TiO2. Spectrophotometric techniques and isothermal titration calorimetry were used to determine the speciation of Ti(IV)-DFOB and revealed a stability constant of log ~ 40 for Ti(IV)-DFOB when in competition with EDTA. Complementary computational methods were employed to predict the structure of Ti(IV)-DFOB, because no crystal structure has been determined thus far. Dissolution studies of TiO2 in the presence of DFOB were monitored by UV/Vis and ICP-OES to determine the kinetics of Ti(IV)-DFOB formation, using many different crystalline forms of TiO2 at several pH values. Kinetic data confirmed that dissolution of Ti(IV) with DFOB is a two-step process, with one faster, less extensive step and a slower step involving additional Ti(IV). Introduction of small organic acid-derived ligands such as oxalate, citrate, ascorbate and succinate changed the dissolution kinetics, suggesting a synergistic cooperation between oxalate-DFOB dissolution, while the others revealed inhibitory behavior. Exposure of sunscreen products that contained TiO2 to DFOB was also investigated to determine biological effects on siderophore binding. Further investigative studies were conducted using SEM and TEM to address the surface interactions of TiO2 with DFOB. Understanding these interactions is necessary to determine the effects of binding, the interactions of these complexes in aqueous environments and how they behave chemically in biological systems. Varying concentrations of Fe(III) and Ti(IV) were introduced together with DFOB to determine by using UV/Vis spectroscopy what metal will bind preferentially. ESI-mass spectra were obtained of these solutions to further confirm metal binding. DFOB-mediated mineral dissolution studies were explored by spectrophotometry and ICP-OES to determine the amount of soluble metal released into solution from -hematite Fe2O3, anatase TiO2 and pseudobrookite (Fe2TiO5) and the kinetics of dissolution. Finally, surface analysis was conducted using SEM and TEM to observe the effects of DFOB on the mineral phase. The demonstration that DFOB can bind Ti(IV) and solubilize TiO2 raised the question of whether other siderophores could potentially cause the same effects. Another biologically relevant siderophore is pyoverdine (PVD), found in Pseudomonas bacteria. It has been a strong focus since it was found to have many important roles ranging from virulence, cell to cell signaling, and quorum sensing for biofilm formation. Adhesion of these bacteria is often found on titanium surfaces. Biofilms form on biomedical titanium implants and biologically induced corrosion often occurs on TiO2 coated surfaces, such as on the sides of ships and the interior of pipes. PVD was isolated from bacterial culture and characterized. PVD was then exposed to Ti(IV) solutions and monitored by UV/Vis spectroscopy and fluorescence to characterize Ti-PVD formation and speciation, by using the same techniques as Ti-DFOB. Binding of Ti(IV) to PVD was determined using ITC to have a log ~51. Treatment of TiO2 with PVD yielded different results from those observed with DFOB. In particular, putative adsorption of PVD to the surface was seen rather than dissolution of Ti(IV). Growth of Pseudomonas in the presence of TiO2 showed enhanced growth rates and using Ti(IV) complexes, the effects on biofilm growth were determined. Understanding these interactions is necessary to determine the effects of binding, the interaction of these complexes in aqueous environments and how they behave chemically in biological systems.