Characterization of the activity of the different oligomeric forms of pulmonary human surfactant protein SP-D

Surfactant protein SP-D is a collectin protein found in mucosal tissues, including lungs. SP-D participates in the innate immune defense of the lungs, where it recognizes and binds pathogens, such as bacteria and viruses, to promote their clearance. SP-D can be assembled into different oligomeric forms: trimers, hexamers, dodecamers and large oligomers. Currently, it is unknown the precise quantitative distribution of the different oligomeric forms in native human SP-D, whether they are all functional structures and, if this is not the case, which is the most active form. We hypothesize that dodecamers and large oligomeric forms might be the most abundant and active structures in human SP-D. In the present work, we have used atomic force microscopy to analyze the structure and quantify the different oligomeric forms and compared the activity of native human SP-D from two different natural sources, amniotic fluid and bronchoalveolar lavage of proteinosis patients. In addition, we have produced a recombinant human version of the protein in CHO cells and isolated the different oligomers to assess their differences in functional behavior. Functional assays, testing the ability of the different protein oligomers to bind and aggregate bacteria, have shown that large oligomers are the most active oligomeric form, followed by hexamers. Trimers bind to bacteria, but they are not able to induce their aggregation. These results suggest that human SP-D from proteinosis patient’s has higher aggregation activity than SP-D from amniotic fluid. Higher activity correlates with a significantly higher proportion of large oligomers in proteinosis SP-D than the protein form amniotic fluid.