FcRn Binds and Transports Albumin

描述(由申请人提供)：我们测试了一个被遗忘了40年的假设，即与MHC相关的Fc受体Ig G(FcRN)，Brambell受体保护白蛋白免受细胞内分解代谢降解，就像它做Ig一样，延长了两个分子的半衰期。我们的初步研究证实了这一假说的三个预测，表明白蛋白在低pH值下与FcRN结合，就像IgG一样，在不同于IgG结合位点的位置形成三分子复合体。初步实验表明白蛋白直接与天然受体结合；SPR实验表明Kd在FM范围内。此外，我们发现FcRN缺陷(KO)小鼠比WT小鼠更快地降解白蛋白，并且FcRN缺陷小鼠的血清白蛋白浓度大约是WT对照小鼠的一半。这些数据推断，在单位时间内，更多的白蛋白分子被保护而不被降解，FcRN可能以某种方式参与了白蛋白的生物合成。 我们的具体目标有两个方面。首先，我们将通过测定相互作用的亲和力、动力学速率常数、特异性和化学计量来表征白蛋白与FcRN的分子相互作用。其次，我们将检验这一假设的六个预测或扩展；即，来自组织的天然FcRN将结合白蛋白，白蛋白可能从母亲转运到后代，FcRN介导白蛋白的跨内皮细胞流动，FcRN参与肝细胞白蛋白的产生，白蛋白在体外通过细胞的运输是FcRN介导的，以及由于白蛋白和免疫球蛋白分子的加速降解速度而缺乏白蛋白和IgG的家庭具有缺陷的FcRN基因。 我们应用了广泛的方法，涵盖了许多学科。在第一个目标中，主要是生物化学，我们使用了表面等离子激元共振，放射性配基结合分析，尺寸排除层析，等温滴定量热法和亲和层析。第二个目标包括几个生理目标，包括药代动力学分析、KO小鼠体内转运研究、组织培养、免疫荧光和分子生物学。 这些研究的影响涉及广泛的健康和疾病主题。这些包括血清白蛋白浓度的动态平衡调节；白蛋白在血浆和血管外室之间的跨内皮流量以及跨其他组织屏障的流量；肝细胞白蛋白的产生机制；蛋白质的胎盘转运；以及MHC蛋白家族的进化。

DESCRIPTION (provided by applicant): We tested a forgotten 40-year old hypothesis that the Brambell receptor, the MHC-related Fc receptor for IgG (FcRn), protects albumin from intracellular catabolic degradation just is it does IgG, prolonging the half-lives of both molecules. Our preliminary studies affirmed three predictions of this hypothesis, showing that albumin binds FcRn at low pH, just as does IgG, at a site distinct from the IgG binding site, forming a trimolecular complex. Preliminary experiments indicate direct binding of albumin to the native receptor; SPR experiments show KD in fM range. Further, we found that FcRn-deficient (KO) mice degrade albumin more rapidly than WT mice, and that the serum albumin concentration in FcRn-deficient mice is about half that of WT controls. These data infer that many fold more albumin molecules are protected from degradation per IgG molecule per unit time, and that FcRn may somehow be involved in albumin biosynthesis. Our specific aims are two fold. First, we will characterize the molecular interaction of albumin with FcRn by determining affinity, kinetic rate constants, specificity, and stoichiometry of interaction. Second, we will test six predictions or extensions of the hypothesis; namely, that native FcRn from tissue will bind albumin, that albumin may be transported from mother to offspring, that FcRn mediates trans-endothelial flux of albumin, that FcRn participates in hepatocyte production of albumin, that albumin transport across cell in vitro is FcRn mediated, and that a family deficient in albumin and IgG because of accelerated degradation rates of both molecules have a faulty FcRn gene. We apply a broad range of methods encompassing many disciplines. In the first aim, mostly biochemical, we use surface plasmon resonance, radioligand binding assays, size exclusion chromatography, isothermal titration calorimetry, and affinity chromatography. The second aim, which includes several physiological goals, employs pharmacokinetic analysis, in vivo transport studies in KO mice, tissue culture, immunofluorescence, and molecular biology. The implications of these studies extend far on a broad range of health and disease topics. These include the homeostatic regulation of serum albumin concentration; the flux of albumin across endothelium between plasma and extravascular compartments arid across other tissue barriers; the mechanism of hepatocyte albumin production; the placental transport of proteins; and the evolution of the MHC family of proteins.