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X-ray crystallography has been used to identify a novel enzyme that polymerises the essential biochemical building block phosphate in eukaryotes, which include all animals, plants, fungi, and protists. The proof of principle was obtained with yeast and could pave the way to the discovery of related enzymes in other species. Phosphate is a ubiquitous multifunctional building block in biochemical energy storage, although in its polyphosphate form it has countless other functions, such as stress response and bone calcification. Scientists have long known how bacteria produce phosphate chains but understanding how this process works in eukaryotes has remained a puzzle. Until now. Researchers at the European Molecular Biology Laboratory (EMBL) in Heidelberg, Germany, and colleagues have now revealed how lower eukaryotes, organisms whose cells have a nucleus, including some fungi and protozoan parasites, polymerise phosphate enzymatically. EMBL's Klaus Scheffzek and colleagues there and at the Department of Biochemistry at the University of Lausanne, Switzerland, and Ruhr University in Bochum, Germany, published details in a recent issue of Science. In their paper, the researchers reveal the function of a single protein domain that might have a wide range of potential applications ranging from improving crops to fighting diseases such as sleeping sickness and Chagas disease. The team obtained a 2.6 ångstrom crystal structure of the catalytic domain grown in the presence of ATP, which they explain revealed polyphosphate winding through a tunnel-shaped pocket. "Nucleotide- and phosphate-bound structures suggest that the enzyme functions by metal-assisted cleavage of the ATP gamma-phosphate, which is then in-line transferred to an acceptor phosphate to form polyphosphate chains," the team explains. In yeast, the protein Vtc4p is part of a protein complex, the vacuolar transporter chaperone complex (VTC). This complex is usually found in the membranes of vacuoles, pockets within which cells store biomolecules for use, transport or destruction. It is is Vtc4p that the team has demonstrated is responsible for the production of polyphosphates. "This protein is like a factory," explains Scheffzek, "it sits in the vacuolar membrane, generates long chains of polyphosphates and we speculate that it sends them straight to the vacuole for storage." Vtc4p is in fact only embedded partially in the membrane and has a "tail" that protrudes into the cell, as revealed by the crystallography. This tail grabs on to a phosphate group from passing, adenosine triphosphate. This releases the energy from the ATP. The energy released is used by Vtc4p to tack the newly acquired phosphate on to a growing phosphate chain. The next step is speculative but it seems a reasonable assumption that because the remainder of the Vtc4p straddles the membrane it presumably transfers the polyphosphate chain into the vacuole itself. "This study emphasises the importance of structural biology not just to show what molecules look like and how they work but also what that function is," explains EMBL team member Michael Hothorn, who is currently at The Salk Institute for Biological Studies, California. The scientists suspect that Vtc4p must play an important role in making phosphate available to plants via the activity of fungi living in the plants' roots. The vacuolar complex can move from the membrane of the vacuole to the cell membrane where it could assemble polyphosphate and transfer it to the exterior of the fungal cell, where it would then be available to the plant. The discovery could therefore have implications for agrochemicals pointing to ways to create novel fertilizers, or enhancers to improve crop yields, given the importance of polyphosphate in plant growth. Similarly, the parasites involved in sleeping sickness and Chagas disease require polyphosphate and so this research might also provide clues to new treatments for these diseases.
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