Structure of a truncated human surfactant protein D is less effective in agglutinating bacteria than the native structure and fails to inhibit haemagglutination by influenza A virus.

Abstract

Surfactant protein D (SP-D) is a lung-specific protein that is synthesized and secreted by lung epithelial cells and is believed to play an important role in lung host defence. This protein belongs to the C-type lectin family, which is characterized by an N-terminal cysteine-rich domain, a collagen-like domain, a neck domain and a carbohydrate recognition domain (CRD). To elucidate the biological actions of this animal lectin against such pathogens as micro-organisms, the biological activities of a recombinant partial SP-D lacking a collagen-like domain were examined. A recombinant human SP-D, consisting of a short collagen region (two repeats of Gly-Xaa-Yaa amino acid sequences), the neck domain and the CRD, was expressed in Escherichia coli. The recombinant SP-D was purified on a nickel column and then on a maltose-agarose column. This protein can form a trimeric structure owing to the neck domain and exhibits sugar-binding activity and specificity similar to those of native human SP-D. The recombinant SP-D caused dose-dependent and calcium-dependent agglutination of E. coli Y1088. The agglutination titre (the concentration required to achieve a 50% decrease in light transmission by agglutination) of recombinant SP-D was approx. 6-fold that of native SP-D. As for conglutination, the recombinant trimeric conglutinin required 8-16-fold higher concentrations than the native counterpart. In haemagglutination inhibition (HI) of influenza A virus, although native and recombinant conglutinin showed similar levels of HI activity, the recombinant SP-D was unable to inhibit haemagglutination, even at a concentration approx. 120-fold that of the native SP-D. The lectin precipitation and lectin blot assays showed that the truncated SP-D could bind to influenza A virus as well as native SP-D did. These results indicate that the agglutination activity of trimeric collectins can be largely retained, and furthermore that the oligomeric structure with several hands at opposite sites can enhance agglutination activity. The difference in HI activity against influenza A virus between native and recombinant SP-D suggests that SP-D uses a different mechanism from that of conglutinin to inhibit viral haemagglutination.