Secondary structure in lung surfactant SP-B peptides: IR and CD studies of bulk and monolayer phases

AbstractPulmonary surfactant protein SP-B is known to facilitate adsorption and spreading of surfactant components across the air/water interface. This property appears essential for in vivo function in the alveolar subphase and at the air/alveolar surface. Three peptides with amino acid sequences based on SP-B containing predicted α-helical regions (SP-B1-20, SP-B9-36A, SP-B40-60A) have been synthesized to probe structure-function relationships and protein-lipid interaction in bulk phase and monolayer environments. IR and CD studies are reported along with traditional surface pressure-molecular area (π-A) isotherms and IR reflection-absorption spectroscopy (IRRAS) investigations conducted at the air/water interface. In bulk phase, helix-promoting environments (methanol and aqueous dispersions of lipid vesicles), SP-B1-20 and SP-B9-36A contained significant amounts of α-helical structure, whereas varying degrees of α-helix, random coil, and β-sheet were observed in aqueous solutions and monolayers. The most striking behavior was observed for SP-B9-36A, which displayed reversible surface pressure-induced β-sheet formation. Bulk phase lipid melting curves and monolayer experiments with peptide-lipid mixtures showed subtle differences in the degree of bulk phase interaction and substantial differences in peptide surface activity. The uniqueness of IRRAS is emphasized as the importance of evaluating secondary structure in both bulk phase and monolayer environments for lung surfactant peptide mimics is demonstrated

Location of Structural Transitions in an Isotopically Labeled Lung Surfactant SP-B Peptide by IRRAS

Pulmonary surfactant, a lipid/protein complex that lines the air/water interface in the mammalian lung, functions to reduce the work of breathing. Surfactant protein B (SP-B) is a small, hydrophobic protein that is an essential component of this mixture. Structure-function relationships of SP-B are currently under investigation as the protein and its peptide analogs are being incorporated into surfactant replacement therapies. Knowledge of the structure of SP-B and its related peptides in bulk and monolayer phases will facilitate the design of later generation therapeutic agents. Prior infrared reflection-absorption spectroscopic studies reported notable, reversible surface pressure-induced antiparallel β-sheet formation in a synthetic peptide derived from human SP-B, residues 9–36 (SP-B(9–36)). In the current work, infrared reflection-absorption spectroscopy is applied in conjunction with isotopic labeling to detect the site and pressure dependence of the conformational change. SP-B(9–36), synthesized with (13)C=O-labeled Ala residues in positions 26, 28, 30, and 32, shifted the β-sheet marker band to ∼1600 cm(−1) and thus immediately identified this structural element within the labeled region. Surface pressure-induced alterations in the relative intensities of Amide I band constituents are interpreted using a semiempirical transition dipole coupling model. In addition, electron micrographs reveal the formation of tubular myelin structures from in vitro preparations using SP-B(9–36) in place of porcine SP-B indicating that the peptide has the potential to mimic this property of the native protein