DIMERIZATION OF A PROTEIN TYROSINE PHOSPHATASE CD45

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. CD45 is a protein tyrosine phosphatase (PTP) prevalent in most of hematopoietic cells. It is a common leukocyte antigen, and involved in triggering auto-immune response. Recent studies have suggested that reactive oxygen species (ROS) are generated during the activation of tyrosine kinase growth factor receptors and by antigen receptors in lymphocytes. Accumulating evidence suggests that PTPs are regulated by such oxidation and one crystal structural study of PTP1b suggests that major conformational changes may occur in response to oxidation. Thus, ROS generated in lymphocytes may oxidize cysteine residues in CD45 and induce conformational changes. We previously demonstrated that the cysteine oxidation indeed induces dimerization of CD45, as evidenced both in the increase in the radius of gyration (33 to 52 ¿) and the two fold increase in the forward scattered intensity. Our recent study focuses on the examination of the crystal structure which is in clear conflict with our dimer activation hypothesis due to the sever spatial hindrance. We suspect that the crystal contact might have forced the molecule to take a non-physiological conformation. We produced a few mutant versions of CD45, one of which is an exact mimic of the crystallized CD45 constract minus the residues that are disordered and thus invisible in the crystal structure, and carried out additional solution scattering measurements. We are in the process of evaluating the possibly for increased flexibility of the domain structure in solution using the new feature in Modeller, a homology and comparative modeling program developed by the Sali group, to remodel atomic coordinates to fit the predicted scattering curve to the experimental data while staying within the electrostatic and steric constraints in hope of obtaining a more physiological structure in solution.