Multiscale Molecular Modeling Studies Of The Dynamics And Catalytic Mechanisms Of Ironii And Zincii Dependent Metalloenzymes PDF Download

Natural metalloenzymes are often the most proficient catalysts in terms of their activity, selectivity, and ability to operate at mild conditions. However, metalloenzymes are occasionally surprising in their choice of catalytic metals, and in their responses to metal substitution. Indeed, from the isolated standpoint of producing the best catalyst, a chemist designing from first principles would likely choose a different metal. Due to competing evolutionary pressures, many natural enzymes may not have evolved to be ideal catalysts and can be improved for the isolated purpose of catalysis in vitro when the competing factors are removed. To improve and, in due course, design metalloenzymes, extensive sampling and proper treatment of the electronic structure of the bound metal(s), is required, while seamlessly merging the required techniques to assess energies and entropies, or their changes, for the entire system. Approaching these challenges with a multi-scale approach, the Alexandrova group has developed an accurate and efficient quantum mechanical/molecular mechanics (QM/MM) hybrid dynamics method to model metalloproteins called quantum mechanics/discrete molecular mechanics (QM/DMD). QM/DMD operates through an iterative scheme between QM and MM machineries. DMD is a flavor of molecular dynamics (MD) that approximates the continuous interaction potentials in classical MD with square-well potentials, course-graining the potentials and overall reducing the number of calculations needed. Due to these discretized potentials, DMD is driven by collision events rather than physical forces as in traditional MM and MD. Therefore, the user saves a tremendous amount of time with DMD by solving ballistic equations of motions rather than Newtonian equations of motions. This fast and efficient hybrid dynamics tools has allowed us to investigate various metal-dependent phenomena in natural metalloenzymes such as: 1) exploring Nature's curious choices for specific metals using two amide hydrolases that contain different metals as a case study, 2) examining protein conformational responses to substrate binding and metal replacement as showcased by the role of a flexible loop [beta]-lactamase in binding antibiotic substrates and 3) investigating how the species of the metal dictates the reaction mechanism in a pair acireductone dioxygenases (ARD/ARD'). Extending outside the realm of naturally occurring enzymes, our tools have the ability to span across formidable challenges such as metalloenzyme design, where stabilization of a transition state of the catalyzed reaction in the specific binding pocket around the metal needs to be achieved. QM/DMD was used in the redesign of a well-studied Zn2+ peptidase, carboxypeptidase A (CPA), an enzyme involved in the breakdown of proteins, with a slight preference for bulky hydrophobic groups. More specifically, the enzyme and substrate system were modified to create specific-specific binding and subsequent experiments proved the mutant to be catalytically active. Additionally, another tool called Eris-QM/DMD was formulated to better gauge the effect of mutation on protein structure during the design process. Eris is a stand-alone package that evaluates protein stability upon mutagenesis. Coupling the software to QM/DMD gives us the distinct advantage of accounting for the effect of the metal during protein alternations. With this diverse set of tools, our future ambitious goals are to install catalytically potent non-physiological metals into proteins. While nature is limited to operating with bio-available elements, some metals such as Ir, Pd, Sc, and Rh, which have been shown to be excellent catalysts, even surpassing physiological metals. If the catalytic activity of these non-physiological metals can be combined with the superb selectivity and mild operational conditions characteristic of proteins, new proficient enzymes may emerge. Another advantage to enzymatic catalysis, done either in vitro, or in vivo, is that it can be cheaper, "greener", and more efficient than synthetic catalysis. An early endeavor in this frontier of metalloenzyme design involves installing Pd2+ into an existing protein scaffold, specifically for intramolecular hydroarylation of C-C triple bonds to form coumarins.