Design and engineering switchable artificial metalloproteins.

Abstract

Life requires stable temperatures and pH to sustain the complex biological machinery within the cell. To accommodate the strict environmental conditions for life to proceed, biological processes are often regulated by conformational changes. The approaches used in our research combine conformational flexibility of periplasmic binding proteins and the knowledge of the well-established molecularly rigid artificial metalloproteins to probe the role and mechanisms of conformational regulation. We are creating switchable artificial metalloproteins (swArMs) that undergo conformational changes upon effector binding at a site allosteric to the installed metallocofactor. Specifically, we have bioconjugated a cobalt complex via cysteine ligation within the glutamine binding protein and observed faster bond breaking upon binding to the allosteric effector, glutamine. We are also working to develop reversible swArMs. To this end, we are characterizing and engineering a periplasmic binding protein (PqqT) that natively binds pyrroloquinoline quinone (PQQ). Our aim is to drive redox changes in PQQ that in turn, drive conformational changes in PqqT. Structural characterization of this association complex through x-ray diffraction is underway that will inform our ability to convert this protein into an artificial methanol dehydrogenase. The described research lays groundwork for studying how protein dynamics influence function, with the goal of understanding what makes metalloproteins in nature so optimal for the chemistry they perform.