May 2003
Volume 44, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2003
The Distribution of Transsulfuration Enzyme, Cystathionine-Beta-Synthase (CBS), in the Eye
Author Affiliations & Notes
  • C. Persa
    Veterinary and Biomedical Sci, University of Nebraska-Lincoln, Lincoln, NE, United States
  • S. Moon
    Veterinary and Biomedical Sci, University of Nebraska-Lincoln, Lincoln, NE, United States
  • M.F. Lou
    Veterinary and Biomedical Sci., and Ophthalmology, University of Nebraska, Lincoln, NE, United States
  • Footnotes
    Commercial Relationships  C. Persa, None; S. Moon, None; M.F. Lou, None.
  • Footnotes
    Support  NIH grant EY 10590
Investigative Ophthalmology & Visual Science May 2003, Vol.44, 422. doi:
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    • Get Citation

      C. Persa, S. Moon, M.F. Lou; The Distribution of Transsulfuration Enzyme, Cystathionine-Beta-Synthase (CBS), in the Eye . Invest. Ophthalmol. Vis. Sci. 2003;44(13):422.

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

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Abstract

Abstract: : Purpose : Clinical abnormalities in cystathionine-beta-synthase (CBS) deficiency, a key enzyme in the transsulfuration pathway, associate with many disordered the eye, frequently with ectopic lentis, myopia and less frequently with retinal degeneration, retinal detachment, optical atrophy, glaucoma, corneal abnormalities and cataracts. However, little is known about this enzyme in the eye. The goal of this study is to examine the distribution of CBS in the various regions of the eye. Methods: Fresh eyes from humans (23-67yrs, Omaha Lions Eye Bank) and pigs (6 months, from Farmland Meat packing Co.) were dissected into conjunctiva, cornea, iris, lens, vitreous, retina and optical nerve. Each tissue was homogenized in lysis buffer and the lysate was used for protein determination, CBS activity assay and western blot analysis. Pure human liver CBS enzyme and its anti-CBS antibody (gifts from R. Banerjee, University of Nebraska) were used as positive controls. The enzyme activity was measured by quantifying the condensation of 14C-U-serine (30mM) with homocysteine (15 mM) to 14C-cystathionine. The reaction mixture of pyridoxal phosphate (0.25 mM), S-adenosylmethionine (0.38 mM) and Tris buffer (pH8.6, 50 mM) in the presence of tissue lysate was incubated at 37oC for 30 min. The reaction was stopped by adding 10% trifluoroacetic acid, and the end product of 14C-cystathionine was separated on a cation exchange column (AG 50W x8) and counted by a scintillation counter. Soluble proteins in tissue homogenate were separated by SDS-PAGE followed by western blot analysis with human liver anti-CBS antibody. Results: Both pig and human eyes showed the highest enzyme activity in cornea and conjunctiva, which was 3-5 X that of iris, vitreous body, retina and optical nerve. Activity in the whole lens was barely detectable. Immunoblotting analysis confirmed the presence of CBS in the above tissues and the pattern of the western blot agreed with that of the enzyme activity. Preliminary data showed that the content of CBS in the epithelial layer of human lens was inversely associated with aging. Conclusion: This is the first evidence of a transsulfuration pathway in the eye. The key enzyme (CBS) is present in all regions of the eye, including vitreous body, but it is more concentrated in the anterior segment and least in the lens. Supported by NIH grant EY 10590. None

Keywords: metabolism • enzymes/enzyme inhibitors • oxidation/oxidative or free radical damage 
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