Glycosylation of a key cubilin Asn residue results in reduced binding to albumin

Shiv Pratap Singh Yadav, Aiying Yu, Jingfu Zhao, Jasdeep Singh, Saloni Kakkar, Srinivas Chakraborty, Yehia Mechref, Bruce Molitoris, Mark C. Wagner

Research output: Contribution to journalArticlepeer-review

Abstract

Kidney disease often manifests with an increase in proteinuria, which can result from both glomerular and/or proximal tubule injury. The proximal tubules are the major site of protein and peptide endocytosis of the glomerular filtrate, and cubilin is the proximal tubule brush border membrane glycoprotein receptor that binds filtered albumin and initiates its processing in proximal tubules. Albumin also undergoes multiple modifications depending upon the physiologic state. We previously documented that carbamylated albumin had reduced cubilin binding, but the effects of cubilin modifications on binding albumin remain unclear. Here, we investigate the cubilin-albumin binding interaction to define the impact of cubilin glycosylation and map the key glycosylation sites while also targeting specific changes in a rat model of proteinuria. We identified a key Asn residue, N1285, that when glycosylated reduced albumin binding. In addition, we found a pH-induced conformation change may contribute to ligand release. To further define the albumin-cubilin binding site, we determined the solution structure of cubilin's albumin-binding domain, CUB7,8, using small-angle X-ray scattering and molecular modeling. We combined this information with mass spectrometry crosslinking experiments of CUB7,8 and albumin that provides a model of the key amino acids required for cubilin-albumin binding. Together, our data supports an important role for glycosylation in regulating the cubilin interaction with albumin, which is altered in proteinuria and provides new insight into the binding interface necessary for the cubilin–albumin interaction.

Original languageEnglish
Article number102371
JournalJournal of Biological Chemistry
Volume298
Issue number10
DOIs
StatePublished - Oct 2022

Keywords

  • albumin
  • cell biology
  • cubilin
  • glycosylation
  • kidney
  • proximal tubule
  • small-angle X-ray scattering (SAXS)

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