BIGH3 WILD TYPE AND MUTANT PROTEINS

已经发现几种角膜营养不良[颗粒状角膜营养不良（GCD）、格子状角膜营养不良（LCD）I型和IIIA型、组合UCD-LCD I型、Thiel-Behnke营养不良和Reis-Bucklers营养不良]由BIGH 3突变引起。这些病症的特征在于与β-Ig-h3（BIGH 3基因的蛋白质产物）的抗体反应的蛋白质在角膜内的沉积。 对于每种营养不良，积聚物具有特征性的超微结构。 该建议是从患有和不患有不同BIGH 3角膜营养不良的人的血浆中以及从COS- 7、CHO-K1和杆状病毒感染的昆虫细胞的细胞培养物中纯化和表征β-Ig-h3，所述昆虫细胞用含有产生BIGH 3角膜营养不良的突变的载体转染。将比较野生型和突变型β-ig-h3蛋白以检验以下假设：在BIGH 3角膜营养不良中在角膜内积累的蛋白的结构与突变蛋白相关，并且不同表型和BIGH 3基因型之间以及表达的突变蛋白的量之间存在关系。将研究在某些BIGH 3角膜营养不良中积累的淀粉样蛋白的性质及其产生的原因。 我们将确定BIGH 3相关淀粉样蛋白是否是突变型β-ig-h3的特异性片段。 因为β-Ig-h3与细胞外基质的组分相互作用，我们将尝试鉴定蛋白质-蛋白质相互作用，其可以解释表征BIGH 3角膜营养不良的积累。将在兔角膜上皮细胞中表征β-Ig-h3的生物合成。

Several corneal dystrophies [granular corneal dystrophy (GCD), lattice corneal dystrophy (LCD) types I and IIIA, combined UCD-LCD type I, Thiel-Behnke dystrophy and Reis-Bucklers dystrophy] have been found to result from mutations in BIGH3. These conditions are characterized by the deposition within the cornea of proteins that react with antibodies to beta-ig-h3 (the protein product of the BIGH3 gene). For each dystrophy the accumulations have a characteristic ultrastructure. This proposal is to purify and characterize beta-ig-h3 from plasma of persons with and without different BIGH3 corneal dystrophies and from cell cultures of COS- 7, CHO-K1 and baculovirus infected insect cells transfected with vectors containing the mutations that produce BIGH3 corneal dystrophies. Wild-type and mutant beta-ig-h3 proteins will be compared to test the hypothesis that the structure of the proteins that accumulate within the cornea in the BIGH3 corneal dystrophies are related to the mutated protein and that a relationship exists between different phenotypes and the BIGH3 genotypes and between the amount of the mutant protein that is expressed. The nature of the amyloid that accumulates in certain BIGH3 corneal dystrophies and the reason for its production will be investigated. We will determine whether BIGH3 related amyloid is a specific fragment of mutant beta-ig-h3. Because beta-ig-h3 interacts with components of the extracellular matrix we will attempt to identify protein- protein interactions that may account for the accumulations that characterize the BIGH3 corneal dystrophies. The biosynthesis of beta-ig-h3 will be characterized in rabbit corneal epithelial cells.