External membrane proteins of rabbit lung macrophages

Abstract

Externally disposed polypeptides of rabbit lung macrophages were labeled using chloramine-T. Optimal conditions, chosen as those which maximized the incorporation of 125I without inhibiting phagocytosis of C3-opsonized lipopolysaccharide oil particles, were found to be dependent on concentrations of carrier iodide, chloramine-T, and the cells themselves. These macrophages inhibit the labeling reaction owing to an apparent abundance of surface sulfhydryl groups which preferentially become oxidized before labeling can occur. Analyzed on polyacrylamide gel electrophoresis, whole macrophages displayed major bands of radioactivity whose apparent molecular weights were: 317,000, 245,000, 186,000, 143,000, and 104,000 daltons. All bands were completely removed by trypsin treatment except a large band of 10,000–15,000 daltons which was removed by lipid solvent extraction and diminished by β-mercaptoethanol treatment of whole labeled cells. No label comigrated with actin at 42,000 daltons or with either of the two major proteins found in the lung lavage fluid. Very similar bands were found in podosomes, peripheral hyaline blebs of plasma membrane, prepared from whole labeled cells.