Isolation of protein components from rat lung lamellar bodies

J. G. Nijssen, D. Hardeman, H. N.M. Lucas Luyckx, L. W. Promes, J. A. Post, H. van den Bosch*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

Lamellar bodies isolated from 10% (w/v) rat lung homogenates by discontinuous sucrose gradient centrifugation were shown to contain variable amounts of adhering proteins. These contaminating proteins could be removed by either Sepharose 4B gel filtration or precipitation of the crude preparation at pH 11.5. Both purification methods yielded membrane preparations with a phospholipid-to-protein ratio of 10.0 μmol/mg. Nearly complete separation of lamellar body phospholipid and protein could be achieved upon application of the purified membranes to DEAE-cellulose in the presence of 0.2% (v/v) Triton X-100. Phospholipid analyses showed that 83% of total lipid phosphorus was recovered in phosphatidylcholine. In phosphatidylethanolamine, phosphatidylglycerol, phosphatidylserine and phosphatidylinositol recoveries amounted to 4, 8, 2 and 2%, respectively. Molecular mass determinations of the isolated protein component of lamellar bodies by means of SDS polyacrylamide gel electrophoresis and staining with Coomassie brilliant blue revealed the presence of three protein bands with molecular masses of 64, 33 and 31 kDa. Upon staining with silver a 16 kDa protein was also visible. Sephadex G-100 gel filtration showed only one protein peak corresponding to a molecular mass of 64 kDa when protein was assayed with Coomassie brilliant blue.

Original languageEnglish
Pages (from-to)131-139
Number of pages9
JournalBiochimica et Biophysica Acta (BBA)/Lipids and Lipid Metabolism
Volume917
Issue number1
DOIs
Publication statusPublished - 13 Jan 1987

Keywords

  • (Rat lung)
  • Lamellar body
  • Protein component

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