Structure, mechanisms, regulation and assembly of ATP synthase

Biological life depends on a source of energy. That energy comes from sunlight. It is harvested by photosynthesis in green plants, and provides the high energy components in the foods that we eat. We harvest that energy in our bodies by "burning" (oxidising) the high energy components, releasing cellular energy in a controlled way to generate the fuel of life, in the form of the molecule known as adenosine triphosphate (or ATP for short). The key steps in this process take place in the mitochondria inside the cells that make up our tissues. The mitochondria are biological "power stations" full of millions of tiny molecular turbines, the ATP synthases, that rotate rather like man-made turbines churning out the cellular fuel in massive quantities, which is then delivered to all parts of our bodies to provide the energy to make them function. We understand in great detail many aspects of how these molecular turbines work, but as yet not how the rotary action is generated. Part of this project is about providing that crucial missing information. Bacteria have similar turbines that differ is some respects from the human turbines. We want to understand these differences better, so that we can devise drugs to kill pathogenic bacteria, such as the one that causes tuberculosis, by stopping their turbines without influencing the human ones. Associated with the human turbines, but not part of the ATP making machinery, is a channel that becomes opened, for example during a heart attack, letting out the contents of the mitochondria in an uncontrolled way leading to the death of the cell. We want to understand how this pore works, so that we can help design other drugs to control the pore.

The F-ATP synthases in eubacteria and mitochondria are rotary machines with many common features. We will build on past achievements to extend fundamental knowledge about them and provide a basis for putting that knowledge to practical use. The mechanism of their catalytic domains is well understood, but how rotation is generated from the transmembrane proton-motive force is less clear. We will provide a molecular explanation of the generation of rotation. We intend to exploit mechanistic differences between human and bacterial enzymes to develop new drugs. One difference concerns their mechanisms of regulation. The mitochondrial enzyme is regulated by the inhibitor protein, IF1; in vitro, it inhibits ATP hydrolysis and not ATP synthesis, but its in vivo role is unclear, except that it inhibits the hydrolase activity of assembly intermediates. It may also inhibit in vivo ATP synthesis by a mechanism that we will investigate possible involving other unidentified proteins. Different regulatory mechanisms operate in eubacteria; for example in Mycobacteria, the enzyme can only make ATP and not hydrolyze it, but we do not yet know how hydrolysis is prevented. Eubacterial and mitochondrial enzymes differ also in their subunit compositions. They have a common core set to allow them to make ATP, but the mammalian mitochondrial enzymes have a "supernumerary" set of seven membrane proteins, each with a single predicted transmembrane alpha-helix. Six of them participate in the formation of a non-specific pore, called the mitochondrial permeability transition pore (PTP) which opens in response to high concentrations of Ca2+ inside mitochondria and leads to cell death, linked to many human diseases. Therefore, we will characterize the structure and properties of the PTP, and develop it as another drug target. As part of a wider collaboration we will study how the human ATP synthase is assembled.

This project will provide fundamental knowledge about one of the key enzymes in biology. This knowledge provides opportunities for commercial exploitation by providing details of a drug target, the ATP synthase, already validated in Mycobacterium tuberculosis, for developing new antibiotics to combat infectious diseases. The permeability transition pore provides an opportunity to develop drugs to control cell death, especially by necrosis. Thus, this research will impact on the academic community by provision of deeper understanding of a fundamental process, and on medicine by deepening our understanding of cell death, and the pharmaceutical industry in the UK and elsewhere by providing new targets for drug development.
Some aspects of the project are technically extremely demanding and at the cutting edge of what is currently possible. Hence, the project provides difficult challenges to be overcome, and new up-to-date techniques to be learned, providing students and post-doctoral scientists with a unique experimental experience, and the opportunity to learn a highly relevant skill-set, thereby enhancing their career prospects.