Four-Step Thermodynamic Model for Membrane Protein Folding

 

If a membrane protein is at thermodynamic equilibrium, one can think of its folding and stability in terms of thermodynamic models that need not mirror the biological assembly process. Such models are nevertheless important for the design of biological experiments because they describe the thermodynamic context within which biological processes must proceed. These processes have evolved to take useful advantage of thermodynamic equilibrium states by either regulating the heights of barriers separating such states or by using metabolic energy to work against them.

Jacobs and White proposed a three-step thermodynamic model for membrane protein folding:

• interfacial partitioning,
• interfacial folding
• insertion

based upon structural and thermodynamic measurements of the partitioning of small hydrophobic peptides and the so-called helical hairpin insertion model. An essential feature of their model, subsequently supported by several theoretical studies , was that the bilayer interface provided a free energy well for initial binding and folding of hydrophobic peptides.

At about the same time, Popot and Engelman proposed a two-stage model for the assembly of alpha-helical proteins in which the helices are first "established" across the membrane and then assemble into functional structures. The idea for this model came from a series of experiments which demonstrated that isolated fragments of bR in lipid bilayers can reassemble spontaneously into a fully functional form, consistent with the native protein residing in a free energy minimum. Combined, these two lines of thought represent a four-step thermodynamic cycle. The four steps, partitioning, folding, insertion, and association, can proceed along an interfacial path, a water path, or a combination of the two. Determination of the free energies for each step along a path allow thermodynamic stabilities to be computed.

The four-step model is useful for both constitutive and non-constitutive membrane proteins. For non-constitutive proteins, the steps progressing from left to right describe the energetics of the natural folding process, while for constitutive proteins, the steps progressing from right to left describe "unfolding". Besides providing a useful thermodynamic scheme, the four-step model summarizes the types of experiments on MP folding that are being pursued in several laboratories.

 

 

 

A four-step thermodynamic cycle for describing the energetics of the partitioning, folding, insertion, and association of an alpha-helix into a fluid lipid bilayer. The process can follow an interfacial path, a water path, or a combination of the two. Studies of folding along the interfacial path are experimentally more tractable.