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Surfactant protein A (SP-A) binds to dipalmitoylphosphatidylcholine (DPPC) and induces phospholipid vesicle aggregation. It also regulates the uptake and secretion of surfactant lipids by alveolar type II cells. We introduced the single mutations Glu195-->Gln (rE195Q), Lys201-->Ala (rK201A) and Lys203-->Ala (rK203A) for rat SP-A, Arg199-->Ala (hR199A) and Lys201-->Ala (hK201A) for human SP-A, and the triple mutations Arg197, Lys201 and Lys203-->Ala (rR197A/K201A/K203A) for rat SP-A, into cDNAs for SP-A, and expressed the recombinant proteins using baculovirus vectors. All recombinant proteins avidly bound to DPPC liposomes. rE195Q, rK201A, rK203A, hR199A and hK201A function with activity comparable to wild type SP-A. Although rR197A/K201A/K203A was a potent inducer of phospholipid vesicle aggregation, it failed to stimulate lipid uptake. rR197A/K201A/K203A was a weak inhibitor for lipid secretion and did not competed with rat [125I]SP-A for receptor occupancy. From these results, we conclude that Lys201 and Lys203 of rat SP-A, and Arg199 and Lys201 of human SP-A are not individually critical for the interaction with lipids and type II cells, and that Glu195 of rat SP-A can be replaced with Gln without loss of SP-A functions. This study also demonstrates that the SP-A-mediated lipid uptake is not directly correlated with phospholipid vesicle aggregation, and that specific interactions of SP-A with type II cells are involved in the lipid uptake process.

Pulmonary surfactant protein A (SP-A) augments the uptake of phospholipid liposomes containing dipalmitoylphosphatidylcholine (DPPC) by alveolar type II cells. The SP-A-mediated uptake process of lipids by type II cells have not been well understood. In the present study we investigated the SP-A-mediated interaction of phospholipids with plasma membrane isolated from alveolar type II cells. SP-A increased the amount of liposomes containing radiolabeled DPPC associated with type II cell plasma membrane by 4-fold compared to the control without SP-A when analyzed by sucrose density gradient centrifugation. This effect is dependent upon the SP-A concentration. The enhancement was inhibited by anti-SP-A antibody and EGTA. When type II cell plasma membrane and liposomes containing [14C]DPPC and [3H]triolein were coincubated with or without SP-A, analysis on sucrose density gradients revealed that the profiles of [14C]DPPC and [3H]triolein in each fraction were almost identical with or without SP-A, indicating that SP-A mediates the binding of liposomes to plasma membrane but not transfer of DPPC. SP-A increased the association of liposomes containing DPPC with the membrane by 2-fold more than that containing 1-palmitoyl-2-linoleoyl-phosphatidylcholine (PLPC). SP-A induced aggregation of phospholipid liposomes containing PLPC as well as those containing DPPC, but the final turbidity of DPPC liposomes aggregated by SP-A was only by 15% greater than that of PLPC liposomes. The amount of DPPC liposomes associated with the plasma membrane derived from type II cells was 2-fold greater than that from liver. We speculate that the SP-A-mediated interaction of lipids with type II cell plasma membrane may contribute, in part, to the lipid uptake process by type II cells.