A novel method of purifying lung surfactant proteins A and D from the lung lavage of alveolar proteinosis patients and from pooled amniotic fluid.
A simple procedure has been developed for the purification of the surfactant proteins SP-A and SP-D from lung lavage of patients with alveolar proteinosis. The SP-D is purified by fractionation of the supernatant obtained after spinning the lavage at 10000 X g for 40 min, while the bulk of the SP-A is purified by fractionation of the pellet. The supernatant is applied to a maltosyl-agarose column and the bound SP-D is specifically eluted using MnCl2. The pellet is solubilised in 6 M urea and, following renaturation, the solubilised proteins are applied to maltosyl-agarose and SP-A eluted using a gradient of EDTA. Both SP-A and SP-D are further purified by gel-filtration on Superose-6. This procedure has also been used to prepare successfully human SP-A and SP-D from amniotic fluid and may be generally applicable to the isolation of these surfactant proteins from lung washings obtained from other species.
Strong
P
P
Department of Biochemistry, University of Oxford, UK.
Kishore
U
U
Morgan
C
C
Lopez Bernal
A
A
Singh
M
M
Reid
K B
KB
D000653:Amniotic Fluid / Q000737:chemistry*; D001232:Aspergillus fumigatus; D001992:Bronchoalveolar Lavage Fluid / Q000737:chemistry*; D002498:Centrifugation; D002712:Chlorides; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002853:Chromatography, Liquid; D004492:Edetic Acid; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000302:isolation & purification*; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000276:immunology; D017895:Manganese Compounds; D011510:Proteolipids / Q000302:isolation & purification* / Q000494:pharmacology; D011649:Pulmonary Alveolar Proteinosis / Q000378:metabolism*; D037662:Pulmonary Surfactant-Associated Protein A; D037663:Pulmonary Surfactant-Associated Protein D; D037661:Pulmonary Surfactant-Associated Proteins; D011663:Pulmonary Surfactants / Q000302:isolation & purification* / Q000494:pharmacology; D012685:Sepharose; D014508:Urea
D002712:Chlorides; D006023:Glycoproteins; D017895:Manganese Compounds; D011510:Proteolipids; D037662:Pulmonary Surfactant-Associated Protein A; D037663:Pulmonary Surfactant-Associated Protein D; D037661:Pulmonary Surfactant-Associated Proteins; D011663:Pulmonary Surfactants; D014508:Urea; D012685:Sepharose; D004492:Edetic Acid; C025340:manganese chloride