Qi Y, Nam K, Spong MC, Banerjee A, Sung R-J, Zhang M, Karplus M, Verdine GL.
Strandwise translocation of a DNA glycosylase on undamaged DNA. Proceedings of the National Academy of Sciences of the United States of America. 2012;109 (4) :1086-1091.
AbstractBase excision repair of genotoxic nucleobase lesions in the genome is critically dependent upon the ability of DNA glycosylases to locate rare sites of damage embedded in a vast excess of undamaged DNA, using only thermal energy to fuel the search process. Considerable interest surrounds the question of how DNA glycosylases translocate efficiently along DNA while maintaining their vigilance for target damaged sites. Here, we report the observation of strandwise translocation of 8-oxoguanine DNA glycosylase, MutM, along undamaged DNA. In these complexes, the protein is obsd. to translocate by one nucleotide on one strand while remaining untranslocated on the complementary strand. We further report that alterations of single base-pairs or a single amino acid substitution (R112A) can induce strandwise translocation. Mol. dynamics simulations confirm that MutM can translocate along DNA in a strandwise fashion. These observations reveal a previously unobserved mode of movement for a DNA-binding protein along the surface of DNA.
Ovchinnikov V, Karplus M.
Analysis and Elimination of a Bias in Targeted Molecular Dynamics Simulations of Conformational Transitions: Application to Calmodulin. J. Phys. Chem. B. 2012;116 (29) :8584-8603.
AbstractThe popular targeted mol. dynamics (TMD) method for generating transition paths in complex biomol. systems is revisited. In a typical TMD transition path, the large-scale changes occur early and the small-scale changes tend to occur later. As a result, the order of events in the computed paths depends on the direction in which the simulations are performed. To identify the origin of this bias, and to propose a method in which the bias is absent, variants of TMD in the restraint formulation are introduced and applied to the complex open ↔ closed transition in the protein calmodulin. Due to the global best-fit rotation that is typically part of the TMD method, the simulated system is guided implicitly along the lowest-frequency normal modes, until the large spatial scales assocd. with these modes are near the target conformation. The remaining portion of the transition is described progressively by higher-frequency modes, which correspond to smaller-scale rearrangements. A straightforward modification of TMD that avoids the global best-fit rotation is the locally restrained TMD (LRTMD) method, in which the biasing potential is constructed from a no. of TMD potentials, each acting on a small connected portion of the protein sequence. With a uniform distribution of these elements, transition paths that lack the length-scale bias are obtained. Trajectories generated by steered MD in dihedral angle space (DSMD), a method that avoids best-fit rotations altogether, also lack the length-scale bias. To examine the importance of the paths generated by TMD, LRTMD, and DSMD in the actual transition, we use the finite-temp. string method to compute the free energy profile assocd. with a transition tube around a path generated by each algorithm. The free energy barriers assocd. with the paths are comparable, suggesting that transitions can occur along each route with similar probabilities. This result indicates that a broad ensemble of paths needs to be calcd. to obtain a full description of conformational changes in biomols. The breadth of the contributing ensemble suggests that energetic barriers for conformational transitions in proteins are offset by entropic contributions that arise from a large no. of possible paths.
Karthikeyan G, Zambaldo C, Barluenga S, Zoete V, Karplus M, Winssinger N.
Asymmetric Synthesis of Pochonin E and F, Revision of Their Proposed Structure, and Their Conversion to Potent Hsp90 Inhibitors. Chemistry - A European Journal. 2012;18 (29) :8978-8986.
AbstractA concise and modular synthesis of pochonin E and F, and their epimers at C-6 established the correct stereochem. of these two natural products to be (6R). Several members of the pochonin family have been shown to bind the heat shock protein 90 (Hsp90), which has been the focus of intense drug discovery efforts. Pochonin E and F as well as their epimers were derivatized into the corresponding pochoximes and further modified at the C-6 position. Mol. dynamics simulations, docking studies, and Hsp90 affinity measurements were performed to evaluate the impact of these modifications.
Crenshaw CM, Nam K, Oo K, Kutchukian PS, Bowman BR, Karplus M, Verdine GL.
Enforced Presentation of an Extrahelical Guanine to the Lesion Recognition Pocket of Human 8-Oxoguanine Glycosylase, hOGG1. Journal of Biological Chemistry. 2012;287 (30) :24916-24928.
AbstractA poorly understood aspect of DNA repair proteins is their ability to identify exceedingly rare sites of damage embedded in a large excess of nearly identical undamaged DNA, while catalyzing repair only at the damaged sites. Progress toward understanding this problem has been made by comparing the structures and biochem. behavior of these enzymes when they are presented with either a target lesion or a corresponding undamaged nucleobase. Trapping and analyzing such DNA-protein complexes is particularly difficult in the case of base extrusion DNA repair proteins because of the complexity of the repair reaction, which involves extrusion of the target base from DNA followed by its insertion into the active site where glycosidic bond cleavage is catalyzed. Here we report the structure of a human 8-oxoguanine (oxoG) DNA glycosylase, hOGG1, in which a normal guanine from DNA has been forcibly inserted into the enzyme active site. Although the interactions of the nucleobase with the active site are only subtly different for G vs. oxoG, hOGG1 fails to catalyze excision of the normal nucleobase. This study demonstrates that even if hOGG1 mistakenly inserts a normal base into its active site, the enzyme can still reject it on the basis of catalytic incompatibility.