April 2009
Volume 50, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2009
Monosaccharide Composition and Asparagine-Linked Oligosaccharide Profile of Interphotoreceptor Retinoid-Binding Protein
Author Affiliations & Notes
  • T. Duncan
    Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, Bethesda, Maryland
  • R. K. Kutty
    Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, Bethesda, Maryland
  • W. Samuel
    Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, Bethesda, Maryland
  • T. M. Redmond
    Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, Bethesda, Maryland
  • Footnotes
    Commercial Relationships  T. Duncan, None; R.K. Kutty, None; W. Samuel, None; T.M. Redmond, None.
  • Footnotes
    Support  Intramural Research Program of the National Institutes of Health, National Eye Institute
Investigative Ophthalmology & Visual Science April 2009, Vol.50, 3001. doi:
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      T. Duncan, R. K. Kutty, W. Samuel, T. M. Redmond; Monosaccharide Composition and Asparagine-Linked Oligosaccharide Profile of Interphotoreceptor Retinoid-Binding Protein. Invest. Ophthalmol. Vis. Sci. 2009;50(13):3001.

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Abstract

Purpose: : Glycosylation is a common post-translational modification of proteins. Glycans are known to play an important role in protein structure and stability which, in turn, may alter the biological function of the protein. Interphotoreceptor retinoid-binding protein (IRBP) is glycosylated and localized to the interphotoreceptor matrix (IPM). A variant form of IRBP, termed IRBP II, has been identified (Wiggert et al. Neurochem Int. 1988; 13:81-87) based on its altered lectin-binding properties. Both forms of IRBP bind visual cycle retinoids and may facilitate the movement of these retinoids across the IPM, however, their precise role in this process remains unclear. The purpose of these experiments was to characterize the monosaccharide composition and the asparagine-linked oligosaccharides of IRBP and its variant form, IRBP II, as a step toward elucidating the functional role of IRBP in retinoid transport across the IPM.

Methods: : The monosaccharide composition of IRBP I and II was determined after acid hydrolysis of protein samples. The released monosaccharides were then analyzed by high pH anion exchange chromatography (HPAEC) with pulsed amperometric detection. The asparagine-linked oligosaccharide profile of IRBP I and II was determined by treating heat-denatured protein samples with trypsin followed by Peptide N-Glycosidase F (PNGase F). The released N-glycans were permethylated and analyzed by MALDI-TOF-MS.

Results: : The monosaccharides identified by HPAEC were the following: fucose, N-acetyl-galactosamine, N-acetyl-glucosamine, galactose, glucose, mannose, N-acetyl-neuraminic acid, and N-glycol-neuraminic acid. The molar percentage of each monosaccharide were similar for both IRBP I and II. The asparagine-linked oligosaccharides of IRBP I and II consisted of both hybrid-type and complex-type glycans. However, the main component of IRBP I was a complex type glycan, whereas that of IRBP II was a hybrid-type structure.

Conclusions: : In general, the monosaccharide composition of IRBP I and II was similar. However, the proportion of each glycan bound to asparagine differed between IRBP I and II. That is, the main asparagine-linked component of IRBP I was a complex type glycan, whereas, that of IRBP II was a hybrid type structure. The structure of the asparagine-linked glycans of IRBP I and II may be important in the possibly differential functions of IRBP I and II in the IPM.

Keywords: retinoids/retinoid binding proteins • glycoconjugates/glycoproteins • protein modifications-post translational 
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