Identification and characterization of receptors for riboflavin carrier protein in the chicken oocyte. Role of the phosphopeptide in mediating receptor interaction

Sooryanarayana, Shubhashish Sarkar, P. Radhakantha Adiga, Sandhya S. Visweswariah

Research output: Contribution to journalArticlepeer-review

Abstract

Riboflavin carrier protein (RCP) is a phosphoglycoprotein found in the egg and the serum of laying birds and other animals. We have investigated the binding of chicken RCP (cRCP) to membranes prepared from the whole chicken oocytes. RCP binding had an absolute requirement for calcium, with an affinity (K(d) 10-8 M) high enough to be physiologically relevant. Ligand blotting experiments using labeled RCP and vitellogenin, with proteins solubilized from oocyte membranes, indicated that RCP and vitellogenin bound specifically to three proteins of Mr 380, 260 and 110 kDa. Vitellogenin also bound to proteins of Mr 515 kDa and 97 kDa, similar in size to those identified by receptor associated protein of RAP. Reduced and carboxyamidated RCP inhibited the binding of 1225I-labeled RCP to chicken oocyte membranes, but recombinant RCP expressed in E. coli, and dephosphorylated RCP, failed to interact with the receptors, indicating that post-translational modifications were necessary for ligand-receptor interaction. The purified phosphopeptide, prepared from tryptic digests of egg white RCP, was able to inhibit the binding of RCP to the receptor proteins, with an affinity comparable to native RCP indicating that the phosphopeptide sequence present in RCP serves as the focal point for RCP-receptor interactions.

Original languageEnglish (US)
Pages (from-to)230-242
Number of pages13
JournalBiochimica et Biophysica Acta - Protein Structure and Molecular Enzymology
Volume1382
Issue number2
DOIs
StatePublished - Feb 17 1998

Keywords

  • Chicken oocyte
  • LDL receptor
  • LRP
  • Riboflavin carrier protein
  • Vitellogenin

ASJC Scopus subject areas

  • Structural Biology
  • Biophysics
  • Biochemistry
  • Molecular Biology

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