January 1995
Volume 36, Issue 1
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Articles  |   January 1995
Rod outer segment-associated N-acetylgalactosaminylphosphotransferase.
Author Affiliations
  • A J Sweatt
    Department of Ophthalmology, Bowman Gray School of Medicine of Wake Forest University, Winston-Salem North Carolina 27157.
  • J Balsamo
    Department of Ophthalmology, Bowman Gray School of Medicine of Wake Forest University, Winston-Salem North Carolina 27157.
  • J Lilien
    Department of Ophthalmology, Bowman Gray School of Medicine of Wake Forest University, Winston-Salem North Carolina 27157.
Investigative Ophthalmology & Visual Science January 1995, Vol.36, 163-173. doi:
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    • Get Citation

      A J Sweatt, J Balsamo, J Lilien; Rod outer segment-associated N-acetylgalactosaminylphosphotransferase.. Invest. Ophthalmol. Vis. Sci. 1995;36(1):163-173.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

PURPOSE: To determine the exact location of a cell surface glycosyltransferase (N-acetylgalactosaminylphosphotransferase, (GalNAcPTase) immunochemically identified in mammalian rod outer segments (ROS), to determine whether anti-GalNAcPTase antibody recognizes retinal molecules that possess transferase activity and to characterize ROS transferase enzyme activity and acceptors. The GalNAcPTase is known to be associated with the adhesion molecule N-cadherin in embryonic avian retinas and with E-cadherin in mammalian pancreatic islet cells. METHODS: Purified, fixed ROS were reacted with anti-chick GalNAcPTase antibody followed by secondary antibody conjugated to colloidal gold and were examined by electron microscopy. Fractions of retinal and ROS proteins enriched in the transferase were obtained through batch adsorption on Sepharose, separated by gel electrophoresis, transferred to nitrocellulose, and either reacted with anti-GalNAcPTase antibody or assayed for transferase activity. Interphotoreceptor matrix (IPM) was examined for the presence of immunoreactive GalNAcPTase by gel electrophoresis and immunoblot. The kinetics and endogenous acceptors of the cow ROS transferase were characterized. RESULTS: ROS are specifically labeled by anti-GalNAcPTase antibody at the cell surface. The immunogold label was associated with the cell surface and with flocculent material adherent to the cell surface. In addition, soluble and particulate fractions of the IPM showed GalNAcPTase-like immunoreactivity. The transferase appears as single immunoreactive band at or near 220 kd. Transferase enzyme activity was present at this position on Western transfers of retinal and ROS proteins. In whole ROS, transferase activity was directed toward endogenous acceptors of very high molecular mass. CONCLUSIONS: The GalNAcPTase is localized on ROS in association with the cell surface and with components of the IPM. The molecule recognized by the anti-GalNAcPTase antibody possesses transferase activity toward itself and a few other proteins, but mostly toward very large molecules that may be IPM proteoglycans. It is not yet known whether the enzyme of the adult retina specifically transfers sugar or sugar-phosphate groups to its acceptors. It is proposed that the ROS GalNAcPTase is involved in the modulation of adhesive phenomena between or within photoreceptors or between photoreceptors and the interphotoreceptor matrix.

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