Research Article| Volume 78, ISSUE 2, P302-308, August 1971

Synthesis of cobalamin coenzymes by human cells in tissue culture

  • Maurice J. Mahoney
    Reprint requests: Dr. Maurice J. Mahoney, Yale University School of Medicine, 333 Cedar St., New Haven, Conn. 06510.
    From the Division of Medical Genetics, Department of Pediatrics and Medicine, Yale University School of Medicine New Haven, Conn., USA
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  • Leon E. Rosenberg
    From the Division of Medical Genetics, Department of Pediatrics and Medicine, Yale University School of Medicine New Haven, Conn., USA
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  • Author Footnotes
    1 M. J. M. was supported by a training grant from the National Institute of Child Health and Human Development (HD 00198).
    2 L. E. R. was supported by a research career development award from the National Institute of Arthritis and Metabolic Diseases (AM 28987).
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      Humans, as well as other mammals, must convert ingested vitamin B12 to active coenzyme forms for biologic activity. We found that human fibroblasts in tissue culture are capable of carrying out this conversion and we report a method to measure the accumulation of newly synthesized coenzymes by the cultured cells. Fibroblasts are grown in culture medium which contains the precursor vitamin, 57Co-hydroxocobalamin. After 4 days the cells are harvested. Cobalamins are extracted from the cells and then separated by thin layer chromatography. In addition to the hydroxocobalamin, two peaks of radioactivity are found which correspond to the coenzyme forms of B12, deoxyadenosylcobalamin and methylcobalamin. The ability to study coenzyme synthesis and accumulation in human cells will enable investigation of normal B12 metabolism and of human disease states where abnormal metabolism is suspected.
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