CRITICAL STRUCTURE-FUNCTION DETERMINANTS WITHIN THE N-TERMINAL REGION OF PULMONARY SURFACTANT PROTEIN SP-B
Alicia G. Serrano 1, Marnie Ryan 2, Timothy E. Weaver 2 and Jesus Perez-Gil 1*
1 Universidad Complutense de Madrid
2 Cincinnati Children's Hospital Medical Center
* To whom correspondence should be addressed. E-mail: perejil{at}bbm1.ucm.es.
Submitted on August 26, 2005
Revised on September 23, 2005
Accepted on 27 September 2005
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Abstract |
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Surfactant protein SP-B is absolutely required for the surface activity of pulmonary surfactant and postnatal lung function. The results of a previous study (Ryan et al., Biochemistry 44, 861-72, 2005) indicated that the N-terminal segment of SP-B, comprising residues 1-9, is specifically required for surface activity, and suggested that prolines 2, 4 and 6 as well as tryptophan 9, may constitute essential structural motifs for protein function. In the present work, we assessed the role of these two motifs in promoting the formation and maintenance of surface active films. Three synthetic peptides were synthesized including a peptide corresponding to the N-terminal 37 amino acids of native SP-B and two variants in which prolines 2, 4, 6 or tryptophan 9 were substituted by alanines. All three synthetic peptides were surface active, as expected from their amphipathic structure. The peptides were also able to insert into dipalmitoylphosphatidylcholine /palmitoyloleoylphosphatidylglycerol (DPPC/POPG, 7:3 w/w ratio) monolayers preformed at pressures higher than 30 mN/m, indicating that they perturb and insert into membranes. Substitution of alanine for tryptophan at position 9 significantly decreased both the rate of adsorption/insertion of the peptide into the interface and re-insertion of surface active material excluded from the film during successive compression-expansion cycles. Substitution of alanines for prolines at positions 2, 4 and 6 did not produce substantial changes in the rate of adsorption/insertion; however re-insertion of surface active material into the expanding interface film was not as effective as in the presence of the native-like peptide. These results suggest that W9 is critical for optimal interface affinity whereas prolines may promote a conformation that facilitates rapid insertion of the peptide into phospholipid monolayers compressed to the highest pressures during compression-expansion cycling.
Key Words:
AIR-LIQUID INTERFACE, LIPID-PROTEIN INTERACTIONS, MONOLAYER, PULMONARY SURFACTANT, SURFACE TENSION
