Characterization of the heparin-binding domain of human extracellular superoxide dismutase.

The C-terminal, heparin-binding domain of human extracellular superoxide dismutase (hEC-SOD) has been studied as a fusion to human carbonic anhydrase II (HCAII). This technique allows the properties of the EC-SOD domain to be characterized. At the same time, it allows us to differentiate the contributions from the domain, from those properties originating from other parts of EC-SOD. The fusion of the 27 C-terminal amino acids of hEC-SOD to the C-terminal of HCAII (FusCC) resulted in the formation of a monomeric protein, which binds to heparin-Sepharose with approximately the same affinity as the tetrameric hEC-SOD. The structure of the fused C-terminal was characterized by CD and NMR spectroscopy and the data were compatible with the presence of alpha-helical structures as suggested by secondary structure predictions. The NMR data show that the C-terminal of FusCC moves independently from the rest of the protein and that its central part is involved in conformational exchange. The NOESY spectra demonstrate that the C-terminal in both FusCC and hEC-SOD binds to heparin, and that arginine side chains take part in the binding.