Mutations in the PARK14 gene are strongly associated with a spectrum of neurological disorders, including Parkinson's disease (PD) and infantile neuroaxonal dystrophy (INAD) through currently unknown mechanisms. The product of the PARK14 gene is an intracellular calcium-independent phospholipase (PLA2G6 or iPLA2?), which has been implicated in numerous cellular pathways. The protein has a unique multi-domain structure and its activity is regulated at several levels. We propose to solve the crystal structure of the protein in order to advance the mechanistic understanding of iPLA2? function in the brain and other organs. An atomic resolution structure of the enzyme is critical for understanding the mechanism of its activity, its regulation and its function in physiological and pathological states. It will provide the foundation for future development of novel therapeutic approaches to treating neurological and cardiovascular diseases as well as diabetes, cancer and muscular dystrophy. iPLA2? modulates membrane properties, produces bioactive lipid messengers such as arachidonic acid and lysophospholipids, and regulates store-operated calcium entry in multiple cell types. Calmodulin (CaM) inhibits iPLA2? enzyme activity in the presence of calcium. Several cofactors reverse the inhibition. Numerous additional regulatory mechanisms have been suggested including oligomerization, ATP binding and interaction with other cofactors and proteins. The large number and variety of signaling pathways affected by iPLA2? and the complexity of its macromolecular interactions complicate defining its function in cellular events and the role it plays in neurological disorders. PARK14 mutations are found in all structural domains of the protein. Studies of these mutations provide a unique opportunity to link regulatory elements and the catalytic activity of iPLA2? to specific signaling mechanisms and functions. Structural information about the conformation of surface epitopes, of the active site and membrane and protein recognition interfaces will be indispensible for these studies. The goal of this proposal is to obtain this structural knowledge to significantly advance the understanding of iPLA2? function. We have crystallized the full-length iPLA2? and now propose to improve the diffraction quality of the crystals and to obtain phasing information to solve the crystal structure of the enzyme. Results from the proposed studies, together with data obtained from biochemical assays, cellular systems and animal models from our group and others will move the entire field forward and will be critical for understanding the functional role of iPLA2? in the brain and other organs including heart and pancreas.
Mutations of the PARK14 gene locus are strongly associated with a spectrum of neurological disorders including Parkinson's disease (PD) and infantile neuroaxonal dystrophy through currently unknown mechanisms. We propose to solve the atomic resolution structure of the enzyme important for understanding the mechanism of its activity, it's regulation and it's function in physiological and pathological states and may enable development of novel therapeutic approaches.
| Malley, Konstantin R; Koroleva, Olga; Miller, Ian et al. (2018) The structure of iPLA2? reveals dimeric active sites and suggests mechanisms of regulation and localization. Nat Commun 9:765 |
