LAR-interacting proteins, or liprins, are a conserved class of neuronal proteins required for proper formation and function of the presynaptic cytomatrix of the active zone (CAZ). Liprins are composed of liprin-a and liprin-(3 subfamilies which physically interact with each other and form protein scaffolds that organize numerous other proteins for the highly regulated release of presynaptic vesicles. Although immunoprecipitation experiments have shown that a and (3 liprins interact across subfamilies via their three C-terminal SAM domains and that they interact within subfamilies through their N-terminal coiled-coil regions, the stoichiometry and gross architecture of their scaffolding structures is unknown. The proposal in this application details experiments to determine what structures liprin SAM and coiled-coil domains form using a combination of biochemical and biophysical studies. Specifically, the SAM and coiled-coil domains of liprin a and (3 will be cloned into distinct plasmids and expressed separately. Initially, the stoichiometry, physical binding constants, and aggregation states will be measured using a combination of equilibrium sedimentation, surface plasmon resonance, and electron microscopy experiments. The determination of a model at atomic resolution of both domains using x-ray crystallography will be a major focus of the project. Such precise structural information should elucidate the conserved scaffolding mechanism of liprins in the CAZ.
Liprin has an undisputed role in the development and maintenance of neural synapses in several organisms and has recently been implicated in the psychological problems of cocaine overdose in humans. Liprin is also expressed in other human tissues besides the brain and scattered reports have linked its a2 and (31 isoforms to cancer. The determination of the scaffolding structure of liprin should provide a basic foundation for the discovery of therapies to treat cocaine overdose victims, cancer, and probably many more mental health problems in humans yet to be associated with liprin.
| Stafford, Ryan L; Tang, Ming-Yun; Sawaya, Michael R et al. (2011) Crystal structure of the central coiled-coil domain from human liprin-ýý2. Biochemistry 50:3807-15 |
| Stafford, Ryan L; Hinde, Elizabeth; Knight, Mary Jane et al. (2011) Tandem SAM domain structure of human Caskin1: a presynaptic, self-assembling scaffold for CASK. Structure 19:1826-36 |
| Stafford, Ryan L; Ear, Jason; Knight, Mary Jane et al. (2011) The molecular basis of the Caskin1 and Mint1 interaction with CASK. J Mol Biol 412:3-13 |
