Curium(III) complexation with surface-layer (S-layer) proteins from a uranium mining waste pile isolate

During the last years the interaction of U(VI) with the isolate Lysinibacillus sphaericus JG-A12, recovered from the uranium mining waste pile “Haberland” near Johanngeorgenstadt (JG), Saxony, Germany, was intensively studied [1, 2]. It could be demonstrated that L. sphaericus JG-A12 shows the ability to form S-layer proteins. This outermost subcellular structure plays an important role for selective binding of heavy metals, for example U(VI) [2]. The interaction of trivalent actinides (e.g., curium) with bacterial S-layer proteins is unknown. This knowledge is essential for the understanding of An(III) interactions at biological solid-water interfaces. Dependent on pH and [Ca] S-layer proteins occur as monomers and polymers in aqueous solution. The main aim of this study is to explore the unknown Cm(III) binding on different S-layers and S-layer protein structures.
The direct speciation technique time-resolved laser-induced fluorescence spectroscopy (TRLFS) has been applied at trace Cm(III) concentrations. Two S-layer suspensions a) Ca-reduced protein containing 195 mmol Ca/mol protein and b) Ca-purified protein were investigated. As a result and illustrated in Figure 1, both S­layers have a high affinity to bind Cm(III) over a broad pH range between 3 and 9. The Ca-reduced S­layer proteins can associate Cm(III) in two coordination environments characterized by individual emission spectra and luminescence lifetimes (e.g., 140 and 300 µs). The results can be interpreted by an intensive Cm(III) interaction with S-layer monomers between pH 3 and 5 with monomer concentrations of 100, 60, and 17 %, respectively; and a dominating interaction with S-layer polymers between pH 6 to 9 with polymer concentrations of 84, 91, 91, and 75 %, respectively. Whereas in the Ca-purified S-layer protein suspension only one Cm(III) coordination environment occurs with a long lifetime of 300 µs. Here the S-layer monomers dominate the whole pH range. The Cm(III)-S-layer species with monomers identified in both protein suspensions is characterized by an intensive and sharp emission band at 602.6 nm. This suggests an incorporation of Cm(III) into the S-layer protein structure. There is high evidence that the main functionality of the S-layer proteins in Cm(III) binding are the carboxyl groups. A Ca titration of the Cm(III) loaded Ca-purified S-layer protein suspension at pH 9 gave evidence for a change of the S­layer speciation probably by forming polymers. This affects the Cm(III) speciation according to the results detected in the Ca-reduced S­layer suspension. In contrast at pH 3 no indications were observed for an coordination change of Cm(III) bound to the Ca-purified S-layer proteins.
This study presents the first speciation results of Cm(III) with S-layer proteins isolated from bacteria recovered from an actinide contaminated environment. This helps to increase our understanding regarding the speciation of Cm(III) at biological solid-water interfaces. The results of the present study are the basis for a detailed investigation of Ca(II) exchange reactions by Cm(III) in proteins in order to understand the important impact of trivalent actinides on the Ca-metabolism in bacterial cells. Also with respect to a possible An(III) transport in living cells.

[1] J. Raff, U. Soltmann, S. Matys, S. Selenska-Pobell, W. Pompe, H. Böttcher (2003). "Biosorption of Uranium and Copper by Biocers." Chem. Mater. 15: 240-244.
[2] M. Merroun, J. Raff, A. Rossberg, C. Hennig, T. Reich, S. Selenska-Pobell (2005). "Complexation of Uranium by Cells and S-layer Sheets of Bacillus sphaericus JG-A12." Appl. Environ. Microbiol. 71: 5532-5543.

The authors are indebted for the use of the 248Cm to the U.S. Department of Energy, Office of Basic Energy Sciences, through the transplutonium element production facilities at Oak Ridge National Laboratory which was made available as part of a collaboration between the HZDR and the Lawrence Berkeley National Laboratory (LBNL).