Low-molecular-weight hydrophobic proteins from bovine pulmonary surfactant.
Pulmonary surfactant stabilizes the lung by reducing the surface tension in the terminal air spaces. Lipid extract surfactant contains approx. 1% (w/w) low-molecular-weight hydrophobic proteins SP-B (15 kDa: nonreduced) and SP-C (3.5 kDa) and with the remainder being mainly phospholipids. The hydrophobic proteins were purified from bovine lipid extract surfactant using delipidation by phospholipase C digestion followed by hydroxyapatite chromatography. The phospholipase C step removed most of phosphatidylcholine resulting in a 10-fold enrichment of hydrophobic proteins relative to phospholipid. Chromatography of this preparation on a hydroxyapatite column resulted in the elution of phospholipids followed by SP-C and then SP-B. The column chromatography was repeated to remove residual phospholipids and yield purified SP-B and SP-C. The final recovery of SP-B from the lipid extracts was about 15-20% and that of SP-C was 5-10%. The bovine surfactant proteins were reconstituted with phospholipids and examined for their ability to lower the surface tension with a pulsating bubble surfactometer. Reconstituted surfactant preparations containing SP-B and dipalmitoylphosphatidylcholine plus dioleoylphosphatidylglycerol were capable of reducing the surface tension to near zero values at minimum bubble radius while the reconstitutes with SP-C only lowered the surface tension to approx. 20 mN/m. A more rapid decrease in surface tension was observed with reconstituted samples containing both hydrophobic proteins. These results indicate that both SP-B and SP-C can promote the adsorption and spreading of surfactant lipids at the air/liquid interface. In addition, SP-B appears to facilitate the squeeze-out of unsaturated phospholipids leading to an enrichment of dipalmitoylphosphatidylcholine in the monolayer.
Mathialagan
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N
MRC Group in Fetal and Neonatal Health & Development, University of Western Ontario, London, Canada.
Possmayer
F
F
D015060:1,2-Dipalmitoylphosphatidylcholine / Q000494:pharmacology; D000818:Animals; D002417:Cattle; D002845:Chromatography; D008970:Molecular Weight; D010715:Phosphatidylglycerols / Q000494:pharmacology; D010743:Phospholipids / Q000032:analysis; D011510:Proteolipids / Q000302:isolation & purification* / Q000494:pharmacology; D011663:Pulmonary Surfactants / Q000032:analysis* / Q000302:isolation & purification* / Q000494:pharmacology; D013500:Surface Tension; D010738:Type C Phospholipases
D010715:Phosphatidylglycerols; D010743:Phospholipids; D011510:Proteolipids; D011663:Pulmonary Surfactants; D015060:1,2-Dipalmitoylphosphatidylcholine; C051388:1,2-dioleoyl-sn-glycero-3-phosphoglycerol; D010738:Type C Phospholipases