The major glycolipid recognized by SP-D in surfactant is phosphatidylinositol.
Surfactant protein D (SP-D), a multimeric calcium-dependent lectin isolated from pulmonary alveolar lavage, has been previously shown to interact reversibly with crude surfactant [Persson et al. (1990) J. Biol. Chem. 265, 5755-5760]. In this study, SP-D is shown to interact reversibly with a preparation of organelles enriched in lamellar bodies, in a manner inhibited by calcium-chelating agents and by competing saccharides. An interaction with an endogenous glycoprotein could not be identified by electrophoresis of surfactant or lamellar body-associated proteins followed by electrotransfer of the separated proteins to nitrocellulose and then probing with radioiodinated SP-D via lectin overlay. Separation of the surfactant or lamellar body lipids on two-dimensional thin-layer chromatography (2D-TLC) followed by probing with radioiodinated SP-D via lectin overlay demonstrated binding to a single lipid. This interaction was dependent on the presence of calcium and was inhibited by competing saccharides. By assaying column fractions for the ability to bind radioiodinated SP-D after TLC, the glycolipid was purified to homogeneity and identified as phosphatidylinositol (PI). Identification was confirmed by mass spectrometry. We further demonstrate the ability of radiolabeled SP-D to bind to PI presented in a lipid bilayer through separation of free SP-D from liposome-bound SP-D on density gradients of Percoll. The interaction of SP-D with PI is dependent on calcium and inhibited by competing saccharides. SP-D binds with similar efficiency to liposomes with mole fractions of PI ranging from 2.5% to 30%, thereby demonstrating the lectin's ability to recognize mole fractions of PI available in surfactant.(ABSTRACT TRUNCATED AT 250 WORDS)
Persson
A V
AV
Department of Medicine, Jewish Hospital, Washington University Medical Center, St. Louis, Missouri 63110.
Gibbons
B J
BJ
Shoemaker
J D
JD
Moxley
M A
MA
Longmore
W J
WJ
D000818:Animals; D002848:Chromatography, DEAE-Cellulose; D002855:Chromatography, Thin Layer; D008401:Gas Chromatography-Mass Spectrometry; D006017:Glycolipids / Q000378:metabolism*; D006023:Glycoproteins / Q000378:metabolism*; D008081:Liposomes; D008297:Male; D010716:Phosphatidylinositols / Q000378:metabolism*; D037663:Pulmonary Surfactant-Associated Protein D; D011663:Pulmonary Surfactants / Q000737:chemistry* / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013379:Substrate Specificity
D006017:Glycolipids; D006023:Glycoproteins; D008081:Liposomes; D010716:Phosphatidylinositols; D037663:Pulmonary Surfactant-Associated Protein D; D011663:Pulmonary Surfactants