Previous studies have shown that pulmonary surfactant protein D (SP-D) is 
composed of a 43-kDa polypeptide with a short NH2-terminal domain, a collagen 
sequence, and a COOH-terminal C-type lectin domain. In the present studies, 
ultrastructural and biochemical techniques were used to examine the quaternary 
structure of native rat SP-D (rSP-D). Electron microscopy of freeze-dried 
preparations demonstrated a highly homogeneous population of molecules with four 
identical rod-like arms (46 nm in length), each with an 8-9-nm diameter globular 
terminal expansion. The arms, which are similar in diameter to the type I 
collagen helix (approximately 4 nm), emanate from the central "hub" in two pairs 
that closely parallel each other for their first 10 nm. This structure is 
consistent with hydrodynamic studies that predict an highly asymmetric and 
extended molecule (f/f0 = 3.26) with a large Stokes radius (Rs = 18 nm). Pepsin 
digestion gave glycosylated, trimeric collagenous fragments (43 +/- 4 nm, 17 
kDa/chain). Trimeric subunits containing intact triple helical domains were also 
liberated from SP-D dodecamers by sulfhydryl reduction under non-denaturing 
conditions. Digestion of rSP-D with bacterial collagenase generated a 
COOH-terminal carbohydrate binding fragment and a smaller peptide (approximately 
12 kDa, unreduced) that contains interchain disulfide bonds. Electron microscopy 
also demonstrated higher orders of multimerization, with as many as 8 molecules 
associated at the hub. These studies demonstrate that SP-D is assembled as 
homopolymers of four identical trimeric subunits, that interactions between the 
amino-terminal domains of the trimers are stabilized by interchain disulfide 
bonds, and that SP-D molecules can associate to form complex multimolecular 
assemblies.