DETERMINATION OF THE STRUCTURAL ROLE OF THE N-TERMINAL DOMAIN OF HUMAN EXTRACELLULAR-SUPEROXIDE DISMUTASE BY USE OF PROTEIN FUSIONS

The N-terminal domain, containing the 49 N-terminal amino-acid residue s, of human extracellular superoxide dismutase (hEC-SOD) has been stud ied after construction of fusion proteins comprised of the defined dom ain and human carbonic anhydrase II (HCAII). The specific advantage of this technique is that it allows characterization of properties that are intrinsic to the N-terminal domain of hEC-SOD, i.e., the results a re not obscured by properties pertaining to the rest of the hEC-SOD mo lecule. Moreover, the fusion to HCAII allows a rapid and gentle one-st ep purification by affinity chromatography. When the N-terminal domain was fused to the N-terminal of HCAII (= FusNN) a well defined structu re was formed and the resulting protein was tetrameric. When the same hEC-SOD-derived domain was fused to the C-terminal of HCAII (= FusNC), no defined structure of the fused domain could be observed, and the r esulting protein was monomeric. It was concluded that a 'free' N-termi nus is required for formation of the proper structure of the N-termina l domain.