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.
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