A TiO2‐Binding Protein isolated from Rhodococcus Strain GIN‐1 (NCIMB 40340) – Purification,Properties and Potential Applications

A Rhodocuccus strain (Rh. GIN1, NCIMB 40340), which is capable of adsorption to titanium dioxide (TiO₂) and TiO₂‐containing coal fly ash particles, has been isolated previously in our Lab. Selectivity experiments showed that the bacterium is capable of adsorption to other metal oxides as well (e.g. magnetite and Al₂O₃) but at lower affinities. The bacterium binds tightly both to rutile and anatase TiO₂. In electronmicrograms the formation of “bridge‐like” structures between the bacterium and the oxide is observed. A specific protein fraction, located on the cell wall of the bacterium was isolated from the bacterium. This protein was found to adhere strongly to TiO₂ particles at high salt concentrations, similarly to the binding to TiO₂ of the intact bacteria. TiO₂ (rutile) was found to bind the protein faster, stronger and at a higher capacity than the anatase isoform. The 55 kDa Ti‐Binding Protein (TiBP) was isolated from the bacteria after homogenization by French Press. It was purified by affinity chromatography on TiO₂ particles, hydrophobic chromatography on a Fractogel‐propyl column and gel filtration on a Superdex G‐200 column. The same protein was isolated from the bacteria by treatment with mutanolysin, an enzyme which is commonly used to retrieve cell‐wall proteins from Gram‐positive bacteria, demonstrating the outer cell location of the protein in Rh. GIN1. TiBP exhibits metal oxide binding selectivity similar to that of the intact bacterium, namely TiO₂>ZnO>Al₂O₃ >Fe₂O₃(magnetite). Hydrophobic forces seem to dominate the interactions of the protein with TiO₂ as its binding capability is greatly enhanced in the presence of high concentrations of NaCl and its desorption requires high concentrations of urea and SDS. These features differentiate TiBP from other proteins known to adsorb TiO₂ (such as hemoglobin, cytochrome c and bovine serum albumin), mainly by weak, charge‐based interactions.