DIMERIZATION OF A PROTEIN TYROSINE PHOSPHATASE CD45

这个子项目是许多利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 CD45是一种蛋白酪氨酸磷酸酶（PTP），广泛存在于造血细胞中。它是一种常见的白细胞抗原，参与触发自身免疫反应。 最近的研究表明，活性氧（ROS）的产生过程中酪氨酸激酶生长因子受体和抗原受体在淋巴细胞。越来越多的证据表明，PTPs受到这种氧化的调节，PTP1b的一项晶体结构研究表明，氧化可能会发生主要的构象变化。因此，在淋巴细胞中产生的ROS可以氧化CD45中的半胱氨酸残基并诱导构象变化。我们先前证明，半胱氨酸氧化确实诱导CD45的二聚化，如回转半径增加（33至52 <$）和前向散射强度增加两倍所证明的。我们最近的研究集中在检查晶体结构，这是在明确的冲突与我们的二聚体激活假说，由于严重的空间位阻。我们怀疑晶体接触可能迫使分子采取非生理构象。我们产生了几个突变版本的CD45，其中之一是一个精确的模拟结晶CD45 concurrently减去的残基是无序的，因此在晶体结构中不可见的，并进行了额外的溶液散射测量。我们正在评估的过程中，可能增加的灵活性的域结构的解决方案中使用的新功能，在Modeller，同源性和比较建模程序开发的Sali组，重塑原子坐标，以适应预测的散射曲线的实验数据，同时保持在静电和空间的约束，希望获得更多的生理结构的解决方案。

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.