Triblock copolymers of the form PEOαPPOβPEOα [where PEO is poly(ethylene oxide) and PPO is poly(propylene oxide)] form stable insoluble monolayers at both the air−water and solid−water interfaces. At the air−water interface, the nominal equilibrium spreading area of copolymers that have the same PPO content, or “core”, increases with increasing PEO content by an amount equivalent to the area of the hydrated ethylene ether subunits. With compression, a PEO-dependent transition from an expanded to a more condensed monolayer occurs at ∼10 dyn·cm-1, presumably because of condensation of PEO segments onto the core and out of the plane of the surface. Consistent with these results and interpretation, the thickness of adsorbed films of copolymers from series that have fixed cores increases approximately linearly with ethylene oxide content. At collapse of copolymer films, the nominal area per moleculeregardless of the number of ether functions in the PEO segmentis that of the hydrated PPO core. On the basis of these data, we propose models for the structure and ordering of triblock copolymers in condensed films, and we relate these models to the gradation in protein resistance observed for such films.

Annexin V belongs to a family of eukaryotic calcium-dependent membrane-binding proteins. The calcium-binding sites at the annexin-membrane interface have been investigated in some detail; however, little is known about the functional roles of highly conserved interfacial residues that do not coordinate calcium themselves. In the present study, the importance of tryptophan 185, and threonine or serine at positions 72, 144, 228, and 303, in rat annexin V is investigated by site-directed mutagenesis, X-ray crystallography, and functional assays. The high-resolution crystal structures of the mutants show that the mutations do not cause structural perturbations of the annexin molecule itself or disappearance of bound calcium ions from calcium-binding sites. The assays indicate that relative to wild-type annexin V, loss of the methyl substituent at position 72 (Thr72-->Ser) has no effect while loss of the hydroxyl group (Thr72-->Ala or Thr72-->Lys) causes reduction of membrane binding. Multiple lysine substitutions (e.g., Thr72,Ser144,Ser228,Ser303-->Lys) have a greater adverse effect than the single lysine mutation, suggesting that in annexin V the introduction of potentially favorable electrostatic interactions between the lysine side chains and the net negatively charged membrane surface is not sufficient to overcome the loss of the hydroxyl side chains. Replacement of the unique tryptophan, Trp185, by alanine similarly decreases membrane binding affinity. Taken together, the data suggest that the side chains mutated in this study contribute to phospholipid binding and participate directly in intermolecular contacts with phospholipid membrane components.

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