Pulmonary surfactant protein A isolation as a by-product of porcine pulmonary surfactant production.
A pulmonary surfactant reduces surface tension at the air/liquid interface of the alveoli and stabilizes alveoli at low lung volumes. Surfactant deficiency and dysfunction were shown to be present in a number of pulmonary diseases, and surfactant replacement therapy is the common clinical conduct. The hydrophilic SP-A (surfactant protein A) is absent when solvent extraction was used during exogenous surfactant production. Addition of SP-A to the surfactant preparation increases the surface activity and completely counteracts inhibition by blood proteins. SP-A recognizes and binds to carbohydrate structures on the surfaces of pathogenic micro-organisms, and acts as opsonins or cross-linking molecules by binding to a variety of cells that participate in the pulmonary immune response. The purification procedure yielded 206 mg of high-purity SP-A/kg of porcine lung, as judged by gel filtration, SDS/PAGE and Western blotting. The electrophoretic profiles obtained showed that pure SP-A consists of proteins of wide molecular mass in the range 26-36 kDa and a dimer in the range 56-60 kDa. The Western-blot results displayed the same band pattern profile after incubating the membrane using a commercially available polyclonal anti-SP-A antibody produced in goat. Gel-filtration experiments confirmed the molecular mass of SP-A in 10 mM NaCl solution. The isolated SP-A showed mannose-binding ability, representative of its functionality.
Kubrusly
Flávia Saldanha
FS
Centro de Biotecnologia, Instituto Butantan, Avenida Vital Brasil 1500, CEP 05503 900, São Paulo, Brazil. kubruslyfs@butantan.gov.br
Iourtov
Dmitri
D
Leme
Ednilse
E
Raw
Isaias
I
D000818:Animals; D002417:Cattle; D002498:Centrifugation / Q000379:methods*; D002846:Chromatography, Affinity / Q000379:methods*; D002852:Chromatography, Ion Exchange / Q000379:methods*; D037662:Pulmonary Surfactant-Associated Protein A / Q000737:chemistry* / Q000302:isolation & purification*; D011663:Pulmonary Surfactants / Q000737:chemistry* / Q000302:isolation & purification*
D037662:Pulmonary Surfactant-Associated Protein A; D011663:Pulmonary Surfactants