|Title||On the folding of a structurally complex protein to its metastable active state|
|Publication Type||Journal Article|
|Year of Publication||2018|
|Authors||Rao V.VHemanth, Gosavi S|
|Journal||Proceedings of the National Academy of Sciences|
|Keywords||alternate folded conformations, kinetic trap, protein folding simulations, reactive center loop, α1-antitrypsin|
Proteins usually fold to their thermodynamically stable states. The serine protease inhibitors (serpins) modulate several biological processes such as blood clotting and inflammation. These proteins are unusual because their functionally active state, and consequently the state that they need to fold to and populate, is significantly less stable than an alternate latent conformation. This metastability facilitates conformational conversion to the inactive state which irreversibly traps the protease. However, it also makes the serpins prone to disease-linked polymerization. Here, we investigate the structural basis for this conformational choice using folding simulations of the serpin, α1-antitrypsin. We find that the structure of the protease-binding reactive center loop is determined early during folding and gates kinetic accessibility to the latent conformation.
For successful protease inhibition, the reactive center loop (RCL) of the two-domain serine protease inhibitor, α1-antitrypsin (α1-AT), needs to remain exposed in a metastable active conformation. The α1-AT RCL is sequestered in a β-sheet in the stable latent conformation. Thus, to be functional, α1-AT must always fold to a metastable conformation while avoiding folding to a stable conformation. We explore the structural basis of this choice using folding simulations of coarse-grained structure-based models of the two α1-AT conformations. Our simulations capture the key features of folding experiments performed on both conformations. The simulations also show that the free energy barrier to fold to the latent conformation is much larger than the barrier to fold to the active conformation. An entropically stabilized on-pathway intermediate lowers the barrier for folding to the active conformation. In this intermediate, the RCL is in an exposed configuration, and only one of the two α1-AT domains is folded. In contrast, early conversion of the RCL into a β-strand increases the coupling between the two α1-AT domains in the transition state and creates a larger barrier for folding to the latent conformation. Thus, unlike what happens in several proteins, where separate regions promote folding and function, the structure of the RCL, formed early during folding, determines both the conformational and the functional fate of α1-AT. Further, the short 12-residue RCL modulates the free energy barrier and the folding cooperativity of the large 370-residue α1-AT. Finally, we suggest experiments to test the predicted folding mechanism for the latent state.
|Short Title||Proc Natl Acad Sci USA|