Function and regulation of P-type ATPase ion pumps

Summary

The project aims to understand at the atomic level how the Na+,K+-ATPase and related ion pumps function and how they are regulated.

Supervisor(s)

Dr Ron Clarke

Research Location

School of Chemistry

Program Type

PHD

Synopsis

Ion-transporting membrane proteins play a decisive role in the mechanism of all cells and in numerous physiological processes, e.g. ATP production, nerve impulse propagation and muscle contraction. A deeper understanding of their mechanisms and regulation can be obtained by the determination of the kinetics of their individual reaction steps. One approach to this research goal is to make use of the electrical current they generate and to apply an electrophysiological method. Another is to convert the electrical voltage that the proteins produce into an optical signal by incorporating a voltage-sensitive dye into the membrane. Voltage-sensitive styrylpyridinium dyes allow a rapid detection (within nanoseconds) of local electric field changes within membranes. Both spectroscopic and electrical detection methods will be applied in this project.A second important aspect of ion pump function is thermodynamics, i.e. the utilisation of energy. To investigate the energy usage of ion pumps the technique of calorimetry will be applied, which allows one to directly measure the release or absorption of heat when an ion pump undergoes a conformational change or substrate binding/release reaction.

The final essential aspect necessary for a complete understanding of ion pump function is structure. Fortunately, over the last 8 years both the Na+,K+-ATPase and its related enzyme, the Ca2+-ATPase, have been crystallized and structures have been determined by x-ray crystallography. In fact, the Ca2+-ATPase has been crystallized in a number of different conformations around its pump cycle. In this project these structures will be used as a template to construct homology models of the Na+,K+-ATPase and then to investigate its substrate binding via theoretical molecular dynamics simulations.    

Additional Information

Techniques to be used in the project:
* Stopped-flow fluorimetry
* Capacitive current biosensor
* Isothermal titration calorimetry
* Protein isolation and purification
* Homology modelling
* Molecular dynamics simulationsInternational collaborators involved in the project:
* Prof. Flemming Cornelius, University of Aarhus, Denmark
* Prof. Hans-Jürgen Apell, University of Constance, Germany
* Dr. Bela Kelety, Iongate, Frankfurt/Main, Germany
* Dr. Francesco Tadini-Buoninsegni, University of Florence, ItalyScholarships are available to high quality students. Most local students in the laboratory are supported by an Australian or University Postgraduate Award and International students by other scholarships.  Please contact me for further details.

Keywords

Heart disease, kidney function, muscle function, nerve function, body temperature, cell volume, ion transport, kinetics, fluorescence, calorimetry, cell membrane, membrane proteins, phospholipids, cholesterol.

Opportunity ID

The opportunity ID for this research opportunity is: 562