A new model to describe extrinsic protein binding to phospholipid membranes of varying composition: application to human coagulation proteins

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Abstract
We introduce here a new model to describe the binding of extrinsic membrane proteins to acidic lipid membranes. In this view, macroscopic binding affinity is determined by two processes: nonspecific adsorption of protein to the membrane surface and association of acidic lipids with specific sites on the bound protein. We apply this model here to compare the binding of human prothrombin and factor X/Xa to phosphatidylglycerol (PG)- and phosphatidylserine (PS)-containing small unilamellar vesicles measured via relative light scattering. This comparison was undertaken because model membranes containing PS are much more effective in supporting thrombin formation than are membranes containing PG. Analysis of binding isotherms in terms of a traditional membrane binding model gave apparent dissociation constants systematically varying from 0.1 to 10 .mu.M over a range of 8-65 mol % negatively charged phospholipid. With our new description of membrane binding, the dependence of binding data on the acidic lipid surface concentration revealed tht only two or three acidic lipid molecules were associated with each surface-bound factor X/Xa or prothrombin molecule. Assuming four independent and equivalent acidic lipid binding sites per protein, it was possible to adjust the values of only the nonspecific adsorption equilibrium constant and the equilibrium constant describing binding of each species of acidic lipid to individual sites on the protein and thereby obtain a good simulation of log-linear binding isotherms for the full range of acidic lipid surface concentrations. The protein-associated binding sites had a greater affinity for PS than for PG; i.e., a lower surface concentration of PS was required to fill the binding sites. The assumption that 20 acidic lipids, i.e., a sufficient number to form a "pool" or "domain", would associate with each bound protein produced a significantly poorer simulation of the experimental binding isotherms. We conclude that the simple model proposed here offers a physically reasonable, alternative description of coagulation protein binding that avoids the apparently inappropriate assumption of protein-induced phospholipid domains. This model also may be applicable to the binding of other similar extrinsic membrane proteins.