Horowitz A D, Baatz J E, Whitsett J A
Division of Pulmonary Biology, Children's Hospital Medical Center, Children's Hospital Research Foundation, Cincinnati, Ohio 45229.
Biochemistry. 1993 Sep 21;32(37):9513-23. doi: 10.1021/bi00088a001.
The self-association of pulmonary surfactant protein SP-C in lipid vesicles was studied using fluorescence energy transfer. Bovine SP-C was labeled with two fluorescent probes, succinimidyl 6-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]hexanoate and eosin isothiocyanate, on the amino terminus of the protein, producing NBD-SP-C and EITC-SP-C, respectively. The N-terminus of SP-C was relatively immobile between 20 and 37 degrees C, as demonstrated by high fluorescence anisotropy of NBD-SP-C and EITC-SP-C. The mobility increased at the transition of the lipid to the fluid phase. Using fluorescence energy transfer, with NBD-SP-C as the donor and EITC-SP-C as the acceptor, a high degree of SP-C/SP-C association was found below 25 degrees C, decreasing to very little self-association above 42 degrees C in 7:1 1,2-dipalmitoylphosphatidylcholine-1,2-dipalmitoylphosphatidylglycerol (DPPC-DPPG) vesicles. The fraction of SP-C aggregated below 37 degrees C in 7:1 DPPC-DPPG was estimated from the observed energy transfer to be more than 70% of total SP-C. In various lipid mixtures, self-association of SP-C was dependent on the presence of at least some gel-phase lipids. In a lipid mixture resembling pulmonary surfactant, gradually increasing self-association was observed below 38 degrees C. The relation of the present data to the state of aggregation of SP-C in pulmonary surfactant is discussed.
