BIGH3 Wild-Type and Mutant Proteins

描述（由申请人提供）：角膜含有丰富的尚未被理解的临床相关蛋白，对保存角膜透明度很重要。TGFBI （BIGH3）基因突变是几种表型不同的遗传性角膜疾病的原因，这些疾病没有明显的非眼部表现。这些疾病包括几种晶格状角膜营养不良和角膜淀粉样病变、颗粒状角膜营养不良、Reis- B b| cklers角膜营养不良、Thiel-Behnke营养不良和几种非典型角膜疾病。特定的临床和组织病理学表型取决于TGFBI的精确突变，但不同表型的分子解释仍有待确定。由TGFBI编码的细胞外转化生长因子β诱导蛋白（TGFBIp）突变在角膜基质中积累，这些疾病显然仅限于角膜。长期目标是了解这种独特蛋白的特性，并研究TGFBI突变患者角膜内积累的特定沉积物的分子机制。本建议的具体目标是：(1)筛查遗传性角膜疾病受试者TGFBI突变；(2)通过激光捕获显微解剖和液相色谱/串联质谱（LC MS/MS）分析手术切除角膜组织中分离的异常沉积物，以确定部分或全部突变的TGFBIp是否在角膜中积累，以及与之密切相关的其他蛋白质。(3)确定纯化的重组野生型和致病TGFBIp的FAS4结构域的生化和生物物理性质；(4)利用x射线晶体学和核磁共振（NMR）技术在原子分辨率下求解重组野生型TGFBIp的三维结构以及重组野生型和致病突变体的FAS4结构域。公共卫生相关性。这是一项对一种重要的知之甚少的蛋白（TGFBIp）的研究。编码该蛋白的基因（TGFBI）的突变导致几种角膜疾病（营养不良），更好地了解TGFBIp将导致更好的方法来治疗由此导致的视力受损和衰弱症状。

DESCRIPTION (provided by applicant): The cornea contains an abundant poorly understood clinically relevant protein important for the preservation of corneal transparency. Mutations in the TGFBI (BIGH3) gene are responsible for several phenotypically different inherited corneal diseases that have no apparent non-ocular manifestations. These disorders include several varieties of lattice corneal dystrophy and corneal amyloidoses, granular corneal dystrophy, Reis- B|cklers corneal dystrophy, Thiel-Behnke dystrophy, and several atypical corneal disorders. The particular clinical and histopathologic phenotypes are dependent upon the precise mutation in TGFBI, but a molecular explanation for the different phenotypes remains to be determined. The mutated extracellular transforming growth factor beta induced protein (TGFBIp) encoded by TGFBI accumulates in the corneal stroma in these disorders which are apparently limited to the cornea. The long-term objectives are to understand the properties of this unique protein and to investigate the molecular mechanisms responsible for the specific deposits that accumulate within the cornea in patients with mutated TGFBI. The Specific Aims of this proposal are: (1) to screen subjects with inherited corneal diseases for TGFBI mutations, (2) to analyze abnormal deposits isolated from surgically excised corneal tissue by laser capture micro-dissection and liquid chromatography/tandem mass spectrometry (LC MS/MS) to determine whether part or all of the mutated TGFBIp accumulates in the cornea and what other protein(s) are closely linked to it, (3) to determine the biochemical and biophysical properties of the FAS4 domain of purified recombinant wild-type and disease producing TGFBIp and (4) to solve the three-dimensional structure at atomic resolution of recombinant wild- type TGFBIp and the FAS4 domains of recombinant wild-type and disease producing mutants using X-ray crystallography and nuclear magnetic resonance spectroscopy (NMR). PUBLIC HEALTH RELEVANCE. This is a study of an important poorly understood protein (TGFBIp). Mutations in the gene (TGFBI) encoding for this protein cause several corneal diseases (dystrophies) and a better understanding of TGFBIp will lead to better methods of treating the resulting impaired vision and debilitating symptoms.