TY - JOUR
T1 - Mechanism of how carbamylation reduces albumin binding to FcRn contributing to increased vascular clearance
AU - Yadav, Shiv Pratap S.
AU - Sandoval, Ruben M.
AU - Zhao, Jingfu
AU - Huang, Yifan
AU - Wang, Exing
AU - Kumar, Sudhanshu
AU - Campos-Bilderback, Silvia B.
AU - Rhodes, George
AU - Mechref, Yehia
AU - Molitoris, Bruce A.
AU - Wagner, Mark C.
N1 - Publisher Copyright:
Copyright © 2021 the American Physiological Society
PY - 2021/1
Y1 - 2021/1
N2 - Chronic kidney disease results in high serum urea concentrations leading to excessive protein carbamylation, primarily albumin. This is associated with increased cardiovascular disease and mortality. Multiple methods were used to address whether carbamylation alters albumin metabolism. Intravital two-photon imaging of the Munich Wistar Fromter (MWF) rat kidney and liver allowed us to characterize filtration and proximal tubule uptake and liver uptake. Microscale thermophoresis enabled quantification of cubilin (CUB7,8 domain) and FcRn binding. Finally, multiple biophysical methods including dynamic light scattering, small-angle X-ray scattering, LC-MS/MS and in silico analyses were used to identify the critical structural alterations and amino acid modifications of rat albumin. Carbamylation of albumin reduced binding to CUB7,8 and FcRn in a dose-dependent fashion. Carbamylation markedly increased vascular clearance of carbamylated rat serum albumin (cRSA) and altered distribution of cRSA in both the kidney and liver at 16 h post intravenous injection. By evaluating the time course of carbamylation and associated charge, size, shape, and binding parameters in combination with in silico analysis and mass spectrometry, the critical binding interaction impacting carbamylated albumin's reduced FcRn binding was identified as K524. Carbamylation of RSA had no effect on glomerular filtration or proximal tubule uptake. These data indicate urea-mediated time-dependent carbamylation of albumin lysine K524 resulted in reduced binding to CUB7,8 and FcRn that contribute to altered albumin transport, leading to increased vascular clearance and increased liver and endothelial tissue accumulation.
AB - Chronic kidney disease results in high serum urea concentrations leading to excessive protein carbamylation, primarily albumin. This is associated with increased cardiovascular disease and mortality. Multiple methods were used to address whether carbamylation alters albumin metabolism. Intravital two-photon imaging of the Munich Wistar Fromter (MWF) rat kidney and liver allowed us to characterize filtration and proximal tubule uptake and liver uptake. Microscale thermophoresis enabled quantification of cubilin (CUB7,8 domain) and FcRn binding. Finally, multiple biophysical methods including dynamic light scattering, small-angle X-ray scattering, LC-MS/MS and in silico analyses were used to identify the critical structural alterations and amino acid modifications of rat albumin. Carbamylation of albumin reduced binding to CUB7,8 and FcRn in a dose-dependent fashion. Carbamylation markedly increased vascular clearance of carbamylated rat serum albumin (cRSA) and altered distribution of cRSA in both the kidney and liver at 16 h post intravenous injection. By evaluating the time course of carbamylation and associated charge, size, shape, and binding parameters in combination with in silico analysis and mass spectrometry, the critical binding interaction impacting carbamylated albumin's reduced FcRn binding was identified as K524. Carbamylation of RSA had no effect on glomerular filtration or proximal tubule uptake. These data indicate urea-mediated time-dependent carbamylation of albumin lysine K524 resulted in reduced binding to CUB7,8 and FcRn that contribute to altered albumin transport, leading to increased vascular clearance and increased liver and endothelial tissue accumulation.
KW - Albumin
KW - Carbamylation
KW - Cubilin
KW - FcRn
KW - Intravital microscopy
KW - Protein charge
KW - Proximal tubule
KW - Small angle X-ray Scattering
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U2 - 10.1152/AJPRENAL.00428.2020
DO - 10.1152/AJPRENAL.00428.2020
M3 - Article
C2 - 33283642
AN - SCOPUS:85100070195
SN - 1931-857X
VL - 320
SP - F114-F129
JO - American Journal of Physiology - Renal Physiology
JF - American Journal of Physiology - Renal Physiology
IS - 1
ER -