Expression and characterization of the fourth repeat of xenopus interphotoreceptor retinoid-binding protein in E. coli

Claxton A. Baer, Karen L. Kittredge, Alexandra L. Klinger, Deborah M. Briercheck, Mark S Braiman, Federico Gonzalez-Fernandez

Research output: Contribution to journalArticle

15 Scopus citations


Interphotoreceptor retinoid-binding protein (IRBP) is an extracellular glycolipoprotein which in higher vertebrates has a 4-repeat structure and carries endogenous vitamin A and fatty acids. The location of IRBP's 1-2 binding sites for retinol is unknown. To begin to understand which repeat(s) are responsible for ligand-binding, we expressed the fourth repeat of Xenopus IRBP in E. coli to determine if it could by itself bind all-trans retinol. Our expression studies used a polyhistidine fusion domain to purify the recombinant protein directly from inclusion bodies. The fusion protein could be renatured without aggregation if refolded at a sufficiently dilute concentration (< 3 μM). The recombinant fourth repeat of Xenopus IRBP binds [3H]all-trans retinol and the fluorescence of this ligand increases 8-fold upon binding. The binding is saturable with a Kd = 0.4 μM. The expression of recombinant IRBP fragments as fusion proteins in prokayrotes will be useful for defining the structural requirements for ligand binding by this interesting protein.

Original languageEnglish (US)
Pages (from-to)391-400
Number of pages10
JournalCurrent Eye Research
Issue number6
StatePublished - 1994
Externally publishedYes



  • Interphotoreceptor retinoid-binding protein
  • Ligand binding
  • Retina
  • Retinoids
  • Visual cycle
  • Vitamin A-binding proteins
  • Xenopus

ASJC Scopus subject areas

  • Cellular and Molecular Neuroscience
  • Ophthalmology
  • Sensory Systems

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