Kinetic and thermodynamic models for the formation of biomolecular complexes

The formation of noncovalent complexes is an important feature of biological systems in terms of both structure and function. One important biomolecular complex comprises class II proteins of the major histocompatibility complex bound to antigenic peptide fragments. Formation of a kinetic intermediate is an important, although not necessarily universal, aspect of the dynamics of this complex. This process is investigated here by modification of an antigenic peptide to produce a kinetic intermediate, as resolved by high performance size exclusion chromatography. Analysis of a proposed mechanism for the interaction between class II MHC proteins and their peptide ligands shows that dissociation measurements may be ambiguous, and places limits on the range of kinetic parameters that are consistent with a particular set of dissociation experiments. Another important noncovalent interaction in biological mixtures is that between cholesterol and phospholipids in membranes. Numerous unusual thermodynamic properties of monolayer mixtures of cholesterol and phospholipids at the air-water interface have been documented, including the formation of "condensed complexes." In bilayer mixtures, extensive nonideality has been reported, and in cell membranes, lipid microdomains or "rafts" may serve important structural and functional roles. Using regular solution theory, the "average lipid" model of phase behavior in multicomponent cholesterol-phospholipid mixtures is refined. Miscibility critical pressures in ternary cholesterol-phospholipid monolayer mixtures are also studied, and trends therein are explained on the basis of a postulated attractive interaction between phospholipids having dissimilar acyl chains. Properties of hypothetical binary mixtures of cholesterol and phospholipids that form several different condensed complexes are calculated, and it is demonstrated that such mixtures might appear to contain only one complex. Epifluorescence microscopy measurements of phase behavior in ternary mixtures containing molecules associated with membrane "rafts" are used to bridge the gap between monolayer and bilayer studies. The relationship between the chemical activity of cholesterol and its biosynthetic regulation in cells is discussed. An additional connection between monolayer and bilayer experiments is provided by applying the condensed complex model to differential scanning calorimetry experiments performed on cholesterol-phospholipid bilayers. Finally, the condensed complex model is extended to incorporate rate complexes of variable size.