This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.Switching of signals at the surface of plant cells controls plant development, hormone response, avoidance of self-pollination, and disease resistance. Transmembrane signaling in plants is exemplified by the interactions of the receptor-like kinase (RLK) CLAVATA1 with kinase-associated protein phosphatase (KAPP). This pair interacts on the inner surface of the plasma membrane to guide the development of the shoot and flower meristems of the model plant Arabidopsis. The fundamental question of how the KAPP FHA domain interacts with an RLK kinase domain will be investigated. The FHA domain is a newly described class of phosphoserine / phosphothreonine-binding domain which is ubiquitous among eukaryotes in diverse protein-protein signaling contexts. No 3D atomic structures of KAPP or of an RLK have been reported. A high quality structure of the distinctive FHA domain of KAPP, as well as its backbone flexibility, will be determined by NMR for the domain both free and bound to a phosphopeptide. Sequence determinants of the specificity of phosphopeptide interactions with the KAPP FHA domain will be explored using site-directed mutagenesis, screening of phosphoser/thr peptide binding, and thermodynamics of the binding of phosphopeptides most avid for the FHA domain. To further probe how KAPP interacts with CLAVATA1, the structure of the serine kinase domain of this RLK will be modeled. This project will provide insights into how a class of signaling proteins in plants interact specifically during the process of signal switching. It will also add perspective to FHA-mediated protein-protein interactions in other organisms.
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