Calcium dependent association of surfactant protein A with pulmonary surfactant: application to simple surfactant protein A purification

Arch Biochem Biophys. 1996 Mar 15;327(2):285-91. doi: 10.1006/abbi.1996.0123.

Abstract

Surfactant protein A (SP-A) is an abundant lipoprotein component of pulmonary surfactant that plays multiple roles in surfactant homeostasis within the lung. A simple and rapid purification procedure for SP-A is described. Purified surfactant is washed by centrifugation with Ca2+ containing buffer to remove residual soluble proteins. Following the Ca2+ buffer wash, the surfactant pellet is washed in buffer containing EGTA and Mg2+ which releases the bound SP-A in almost pure form. Subsequent chromatography of the SP-A on Sephacryl S-500 yields homogeneous preparations of the protein. The SP-A purified using this procedure requires no exposure to either detergents or organic solvents to remove lipid. SP-A prepared by this new method inhibits lipid secretion from alveolar type II cells as effectively as SP-A prepared by other methods. In addition, the SP-A depleted surfactant produced in the first step of this procedure is capable of binding exogenous SP-A in a time dependent, saturable and Ca2+ dependent manner.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Calcium / pharmacology*
  • Calcium Chloride / pharmacology
  • Chromatography, Gel
  • Egtazic Acid / pharmacology
  • Electrophoresis, Polyacrylamide Gel
  • Enzyme-Linked Immunosorbent Assay
  • Kinetics
  • Lung / metabolism
  • Molecular Weight
  • Protein Binding
  • Proteolipids / drug effects
  • Proteolipids / isolation & purification
  • Proteolipids / metabolism*
  • Pulmonary Alveoli / metabolism
  • Pulmonary Surfactant-Associated Protein A
  • Pulmonary Surfactant-Associated Proteins
  • Pulmonary Surfactants / drug effects
  • Pulmonary Surfactants / isolation & purification
  • Pulmonary Surfactants / metabolism*
  • Rats
  • Rats, Sprague-Dawley

Substances

  • Proteolipids
  • Pulmonary Surfactant-Associated Protein A
  • Pulmonary Surfactant-Associated Proteins
  • Pulmonary Surfactants
  • Egtazic Acid
  • Calcium Chloride
  • Calcium