Prediction of Optimal Folding Routes of Proteins That Satisfy the Principle of Lowest Entropy Loss: Dynamic Contact Maps and Optimal Control

An optimization model is introduced in which proteins try to evade high energy regions of the folding landscape, and prefer low entropy loss routes during folding. We make use of the framework of optimal control whose convenient solution provides practical and useful insight into the sequence of events during folding. We assume that the native state is available. As the protein folds, it makes different set of contacts at different folding steps. The dynamic contact map is constructed from these contacts. The topology of the dynamic contact map changes during the course of folding and this information is utilized in the dynamic optimization model. The solution is obtained using the optimal control theory. We show that the optimal solution can be cast into the form of a Gaussian Network that governs the optimal folding dynamics. Simulation results on three examples (CI2, Sso7d and Villin) show that folding starts by the formation of local clusters. Non-local clusters generally require the formation of several local clusters. Non-local clusters form cooperatively and not sequentially. We also observe that the optimal controller prefers “zipping” or small loop closure steps during folding. The folding routes predicted by the proposed method bear strong resemblance to the results in the literature

Clamp loader ATPases and the evolution of DNA replication machinery

Clamp loaders are pentameric ATPases of the AAA+ family that operate to ensure processive DNA replication. They do so by loading onto DNA the ring-shaped sliding clamps that tether the polymerase to the DNA. Structural and biochemical analysis of clamp loaders has shown how, despite differences in composition across different branches of life, all clamp loaders undergo the same concerted conformational transformations, which generate a binding surface for the open clamp and an internal spiral chamber into which the DNA at the replication fork can slide, triggering ATP hydrolysis, release of the clamp loader, and closure of the clamp round the DNA. We review here the current understanding of the clamp loader mechanism and discuss the implications of the differences between clamp loaders from the different branches of life

Dynamics of open DNA sliding clamps

A range of enzymes in DNA replication and repair bind to DNA-clamps: torus-shaped proteins that encircle double-stranded DNA and act as mobile tethers. Clamps from viruses (such as gp45 from the T4 bacteriophage) and eukaryotes (PCNAs) are homotrimers, each protomer containing two repeats of the DNA-clamp motif, while bacterial clamps (pol III β) are homodimers, each protomer containing three DNA-clamp motifs. Clamps need to be flexible enough to allow opening and loading onto primed DNA by clamp loader complexes. Equilibrium and steered molecular dynamics simulations have been used to study DNA-clamp conformation in open and closed forms. The E. coli and PCNA clamps appear to prefer closed, planar conformations. Remarkably, gp45 appears to prefer an open right-handed spiral conformation in solution, in agreement with previously reported biophysical data. The structural preferences of DNA clamps in solution have implications for understanding the duty cycle of clamp-loaders

Protein folding from a highly disordered denatured state: The folding pathway of chymotrypsin inhibitor 2 at atomic resolution

Previous experimental and theoretical studies have produced high-resolution descriptions of the native and folding transition states of chymotrypsin inhibitor 2 (CI2). In similar fashion, here we use a combination of NMR experiments and molecular dynamics simulations to examine the conformations populated by CI2 in the denatured state. The denatured state is highly unfolded, but there is some residual native helical structure along with hydrophobic clustering in the center of the chain. The lack of persistent nonnative structure in the denatured state reduces barriers that must be overcome, leading to fast folding through a nucleation–condensation mechanism. With the characterization of the denatured state, we have now completed our description of the folding/unfolding pathway of CI2 at atomic resolution

Unusual compactness of a polyproline type II structure

Polyproline type II (PPII) helix has emerged recently as the dominant paradigm for describing the conformation of unfolded polypeptides. However, most experimental observables used to characterize unfolded proteins typically provide only short-range, sequence-local structural information that is both time- and ensemble-averaged, giving limited detail about the long-range structure of the chain. Here, we report a study of a long-range property: the radius of gyration of an alanine-based peptide, Ace-(diaminobutyric acid)(2)-(Ala)(7)-(ornithine)(2)-NH2. This molecule has previously been studied as a model for the unfolded state of proteins under folding conditions and is believed to adopt a PPII fold based on short-range techniques such as NMR and CD. By using synchrotron radiation and small-angle x-ray scattering, we have determined the radius of gyration of this peptide to be 7.4 ± 0.5 Å, which is significantly less than the value expected from an ideal PPII helix in solution (13.1 Å). To further study this contradiction, we have used molecular dynamics simulations using six variants of the AMBER force field and the GROMOS 53A6 force field. However, in all cases, the simulated ensembles underestimate the PPII content while overestimating the experimental radius of gyration. The conformational model that we propose, based on our small angle x-ray scattering results and what is known about this molecule from before, is that of a very flexible, fluctuating structure that on the level of individual residues explores a wide basin around the ideal PPII geometry but is never, or only rarely, in the ideal extended PPII helical conformation

Ca2+ binding by domain 2 plays a critical role in the activation and stabilization of gelsolin

Gelsolin consists of six homologous domains (G1–G6), each containing a conserved Ca-binding site. Occupation of a subset of these sites enables gelsolin to sever and cap actin filaments in a Ca-dependent manner. Here, we present the structures of Ca-free human gelsolin and of Ca-bound human G1–G3 in a complex with actin. These structures closely resemble those determined previously for equine gelsolin. However, the G2 Ca-binding site is occupied in the human G1–G3/actin structure, whereas it is vacant in the equine version. In-depth comparison of the Ca-free and Ca-activated, actin-bound human gelsolin structures suggests G2 and G6 to be cooperative in binding Ca2+ and responsible for opening the G2–G6 latch to expose the F-actin-binding site on G2. Mutational analysis of the G2 and G6 Ca-binding sites demonstrates their interdependence in maintaining the compact structure in the absence of calcium. Examination of Ca binding by G2 in human G1–G3/actin reveals that the Ca2+ locks the G2–G3 interface. Thermal denaturation studies of G2–G3 indicate that Ca binding stabilizes this fragment, driving it into the active conformation. The G2 Ca-binding site is mutated in gelsolin from familial amyloidosis (Finnish-type) patients. This disease initially proceeds through protease cleavage of G2, ultimately to produce a fragment that forms amyloid fibrils. The data presented here support a mechanism whereby the loss of Ca binding by G2 prolongs the lifetime of partially activated, intermediate conformations in which the protease cleavage site is exposed