Mechanistic Investigation into Nitric Oxide Interconversion Pathways of Heme Enzymes using Synthetic Mimics - Project Summary Heme enzymes are powerhouses for catalyzing a plethora of pivotal reaction pathways in humans, and have been the focus of intense research interest throughout the past century. The ubiquitous nature of these enzymes implicates them in a range of physiological functionalities, as well as pathogenic pathways that lead to an array of disease situations including major neurodegenerative and autoimmune conditions, and cancer. In light of that, numerous research efforts have focused on targeting heme enzymes for therapeutic purposes, particularly through meticulously designed selective inhibitors. Nonetheless, a clear mechanistic understanding of a majority of these crucial enzymes is still in its infancy, impeding the rational design of mechanism-based inhibitors that could emerge into promising drug targets. Markedly, most of the mechanistic studies thus far have focused on high-valent heme intermediates, whereby details pertaining to mid-valent heme species (i.e., those containing Fe(III) centers such as heme superoxo, peroxo, hydroperoxo, peroxynitrite, hyponitrite etc.) have remained gravely understudied. Importantly, heme enzymes where mid-valent species are active oxidants and/or key reaction intermediates exhibit a growing significance in human health, making their mechanisms of action of supreme interest. This research program targets this impactful deficit in current knowledge via the utilization of carefully designed synthetic mimic compounds as mechanistic probes. The proposed work is predominantly geared toward gaining a rigorous understanding into heme enzyme mediated nitrogen oxide (NOx/y) interconversion pathways, which, albeit indispensable for normal physiological functions, could lead to nitrative and nitrosative stress in humans under certain conditions. Thus, this project will strive to delineate the exact attributes of heme centers that dictate the geometric and electronic properties of heme-NOx/y intermediates, thereby fine-tuning the structure-activity relationships that choregraph the precision of their biorelevant reactivity pathways. In that, a medley of synthetic heme-NOx/y model complexes supported by systematically varied ligand architectures will be employed (i.e., in relevance to both primary and secondary coordination spheres of the heme center), and their generation, stabilities, and competencies to engage in biorelevant reactivities will be evaluated in detail. Mechanistic investigations will be carried out under cryogenic conditions, which will shine light on the identities and properties of pivotal reaction intermediates; these intermediates will be rigorously analyzed using a combined approach encompassing both spectroscopic and theoretical elements. Moreover, the reaction landscapes of interest will also be examined using exhaustive thermodynamic and kinetic studies; a layer of information that is often exceedingly difficult to obtain solely based on enzymatic systems. This work will therefore further the frontiers of knowledge relevant to NOx/y interconversions facilitated by heme enzymes, which will directly benefit ongoing efforts based on such pathways that target the design and implementation of novel drug candidates with enhanced effectivity and specificity.
