Structural biology of oncogenic receptor tyrosine kinases

描述（由申请人提供）：生长因子通过细胞表面受体酪氨酸激酶（RTKs）信号传导是脊椎动物生物系统中细胞和细胞间调节的最关键途径之一。III类rtk的缺陷/故障，如干细胞因子（SCF）受体KIT、巨噬细胞集落刺激因子（MCSF）受体FMS和FLT3配体（FLT3L）受体FLT3，有助于许多类型癌症的发生和发展，如白血病、胃肠道间质瘤（GIST）等。这些受体是癌症治疗的理想靶点，但其配体识别和激活的机制基础尚不完全清楚。我们的长期目标是阐明这些受体在细胞调控和恶性肿瘤中使用的结构机制。该项目将对III类rtk的整个激活过程进行彻底的分析：当配体不存在时，这些受体如何保持静态状态，它们的细胞外结构域如何识别配体，以及受体激活所需的构象变化是什么。具体目的有：(1)KIT与其配体SCF复合物的生化重构和结构分析。SCF/KIT配合物的晶体衍射x射线到3.2埃的分辨率已经获得。用多波长反常衍射（MAD）来解析其结构。(2) FMS与其配体MCSF配合物的生化重构与结构分析。获得了衍射分辨率为2.4埃的MCSF/FMS配合物晶体。结构由部分分子置换和单同构置换与异常散射（SIRAS）相结合确定；改进正在进行中。(3) FLT3与其配体FLT3L配合物的生化表征及结构分析。最近已经获得了FLT3L/FLT3配合物的小晶体。(4)通过受体/配体界面的蛋白质工程研究受体/配体相互作用的结合特异性和能量学。这些研究将阐明新的结构机制，为治疗iii类rtk相关癌症的治疗开发奠定基础。

DESCRIPTION (provided by applicant): Growth factors signaling through the cell-surface receptor tyrosine kinases (RTKs) represent one of the most critical pathways in the cellular and inter-cellular regulation of vertebrate biological systems. Defects/malfunctions of the class III RTKs, such as the Stem Cell Factor (SCF) receptor KIT, the Macrophage Colony Stimulating Factor (MCSF) receptor FMS, and the FLT3 Ligand (FLT3L) receptor FLT3, contribute to the genesis and development of many types of cancers such as leukemia, gastrointestinal stromal tumor (GIST), and others. These receptors are ideal targets for cancer therapies, but the mechanistic basis underlying their ligand recognition and activation is incompletely understood. Our long-term goal is to elucidate the structural mechanisms used by these receptors in cellular regulation and malignancies. The project will perform a thorough analysis of the whole activation process of class III RTKs: how these receptors maintain static states when the ligands are not present, how their extra cellular domains recognize ligands, and what are the conformational changes required for receptor activation. The specific aims are: (1) Biochemical reconstitution and structural analysis of the complex between KIT and its ligand SCF. Crystals of the SCF/KIT complex that diffract X-rays to 3.2 Angstroms resolution have been obtained. The structure will be solved by multi-wavelength anomalous diffraction (MAD). (2) Biochemical reconstitution and structural analysis of the complex between FMS and its ligand MCSF. Crystals of the MCSF/FMS complex that diffract to 2.4 Angstroms resolution have been obtained. The structure was determined by a combination of partial molecular replacement and single isomorphous replacement with anomalous scattering (SIRAS); refinement is in progress. (3) Biochemical characterization and structural analysis of the complex between FLT3 and its ligand FLT3L. Small crystals of the FLT3L/FLT3 complex have been obtained recently. (4) Investigation of the binding specificity and energetics of the receptor/ligand interactions through protein engineering of the receptor/ligand interfaces. These studies will elucidate novel structural mechanisms that will lay the groundwork for therapeutic development to treat class III-RTK-related cancers.