SP-A binds alpha(1)-antitrypsin in vitro and reduces the association rate constant for neutrophil elastase

BACKGROUND: α1-antitrypsin and surfactant protein-A (SP-A) are major lung defense proteins. With the hypothesis that SP-A could bind α1-antitrypsin, we designed a series of in vitro experiments aimed at investigating the nature and consequences of such an interaction. METHODS AND RESULTS: At an α1-antitrypsin:SP-A molar ratio of 1:1, the interaction resulted in a calcium-dependent decrease of 84.6% in the association rate constant of α1-antitrypsin for neutrophil elastase. The findings were similar when SP-A was coupled with the Z variant of α1-antitrypsin. The carbohydrate recognition domain of SP-A appeared to be a major determinant of the interaction, by recognizing α1-antitrypsin carbohydrate chains. However, binding of SP-A carbohydrate chains to the α1-antitrypsin amino acid backbone and interaction between carbohydrates of both proteins are also possible. Gel filtration chromatography and turnover per inactivation experiments indicated that one part of SP-A binds several molar parts of α1-antitrypsin. CONCLUSION: We conclude that the binding of SP-A to α1-antitrypsin results in a decrease of the inhibition of neutrophil elastase. This interaction could have potential implications in the physiologic regulation of α1-antitrypsin activity, in the pathogenesis of pulmonary emphysema, and in the defense against infectious agents

Study of polytopic membrane protein topological organization as a function of membrane lipid composition.

A protocol is described using lipid mutants and thiol-specific chemical reagents to study lipid-dependent and host-specific membrane protein topogenesis by the substituted-cysteine accessibility method as applied to transmembrane domains (SCAM). SCAM is adapted to follow changes in membrane protein topology as a function of changes in membrane lipid composition. The strategy described can be adapted to any membrane system

Influence of phosphatidylglycerol on the uptake of liposomes by alveolar cells and on lung function

The effect of phosphatidylglycerol on the uptake of surfactant-like liposomes by alveolar type II cells and alveolar macrophages as well as the effect on endogenous surfactant function was studied in vivo. Healthy ventilated rats were intratracheally instilled with fluorescent labeled liposomes with different concentrations of phosphatidylglycerol. Lung function was determined by monitoring arterial oxygenation and, at the end of the experiment, by recording static pressure-volume curves. In addition, alveolar cells were isolated, and cell-associated fluorescence was determined using flow cytometry. The results show that, in the presence of cofactors (Ca2+, Mg 2+), phosphatidylglycerol stimulates the uptake by alveolar macrophages but hardly affects the uptake by alveolar type II cells. High concentrations of phosphatidylglycerol reduce the number of alveolar macrophages in the alveolar space and deteriorate lung function. On the other hand, the presence of cofactors protects the lung against the negative effects of phosphatidylglycerol on endogenous surfactant and alveolar macrophages. This study indicates that the phosphatidylglycerol concentration may play a fundamental role in the surfactant function and metabolism depending on the presence of so-called cofactors like calcium and magnesium; further study is needed to clarify the mechanisms involved