CAREER: Plasticity, Promiscuity and Transport Mechanisms in Transmembrane Metal Pumps

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With the support of the Chemistry of Life Processes Program in the Division of Chemistry, Dr. Gabriele Meloni from The University of Texas at Dallas will study the molecular mechanism of function of transmembrane transporter proteins involved in metal transport across cellular membranes. In all kingdoms of life, the activity of metal pumps guarantee that metal concentrations are regulated to meet cellular requirements. P-type ATPase pumps are a class of transmembrane proteins that utilize ATP as an energy source to move essential and toxic metals across membranes playing a central role in metal balance. However, the general molecular processes involved in specific metal cargo selection and transport remain to be revealed. Structural, biochemical, and biophysical methods will be applied to a comparative characterization of metal recognition in pump subclasses showing diverse metal selectivity patterns to address a new molecular level understanding of metal recognition and translocation. The expected results will contribute to a poorly explored area of biochemistry and will provide new insights regarding metal transport processes in living organisms. The research plan will be integrated with a multifaceted educational program focused on promoting interest and student engagement in bioinorganic chemistry. By developing new course content with peer-led delivery modalities, expanding undergraduate research engagement with a particular focus on underrepresented minorities, and creating a lab-based online video “reality-show” to be shared with Primarily Undergraduate Institutions (PUI) and high school students, the project will promote awareness, interest and societal knowledge in inorganic biochemistry.P1B-type ATPase are ATP-energized primary-active pumps that control translocation of both essential and toxic transition metals. The adaptation of a conserved topological and structural framework, common to all P-type ATPases, to the need of diverse substrate selectivity and promiscuity allowed the evolution of P1B-type subfamilies capable of selective translocation of first-row essential transition metals (Mn(II), Fe(II), Co(II), Ni(II), Cu(I) or Zn(II)), as well as toxic second- and third-row transition and post-transition metals (e.g. Ag(I), Cd(II), Hg(II), Pb(II)). The project will address, by in-house and synchrotron-based biophysical and biochemical methods, the atomic-level determinants controlling metal recognition, promiscuity and structural plasticity in poorly characterized metal pumps subclasses (P1B-4-P1B-7 types) with diverse selectivity patterns. These investigations will be complemented with real-time fluorescence measurements of metal transport events in proteoliposomes combined with the use of fluorescent sensors to investigate putative co-transported ions and membrane potential generation, to provide highlights on the overall kinetics, thermodynamics and mechanism of transport. This integrated approach will reveal new paradigms underlying energy-dependent selective metal translocation processes and define how plasticity and promiscuous recognition is exploited to guarantee metal translocation across membranes.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.