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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1973 Sep;70(9):2585–2589. doi: 10.1073/pnas.70.9.2585

Regulation of 5-Methyltetrahydrofolate:Homocysteine Methyltransferase Activity by Methionine, Vitamin B12, and Folate in Cultured

Daphne Kamely *, John W Littlefield , Richard W Erbe
PMCID: PMC427061  PMID: 4517673

Abstract

Rapid growth of BHK cells in methioninedeficient medium required supplementation with homocysteine, B12, and over 40-fold greater levels of folic acid than growth in methionine-supplemented medium. The activity of the B12-dependent 5-methyltetrahydrofolate: homocysteine methyltransferase was studied in extracts of BHK cells grown in media containing various concentrations of the compounds of the enzyme reaction. The methyltransferase activity increased over 4-fold when B12-deficient deficient medium was supplemented with optimal levels of B12; this increase was not prevented by puromycin. Addition of homocysteine to growth medium containing methionine, B12, and folic acid was without effect. However, methyltransferase activity increased 2.5- to 4.0-fold further beyond the highest levels obtained in the presence of methionine, B12, and folic acid when homocysteine was substituted for methionine in the growth medium. This increase was blocked by puromycin and was not due to removal of feedback inhibition of activity by the product methionine. These results suggest that methyltransferase activity may be regulated in part by derepression of the enzyme's synthesis on substitution of the substrate homocysteine for the product methionine.

Keywords: derepression, cell culture

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. BUCHANAN J. M., ELFORD H. L., LOUGHLIN R. E., MCDOUGALL B. M., ROSENTHAL S. THE ROLE OF VITAMIN B12 IN METHYL TRANSFER TO HOMOCYSTEINE. Ann N Y Acad Sci. 1964 Apr 24;112:756–773. doi: 10.1111/j.1749-6632.1964.tb45053.x. [DOI] [PubMed] [Google Scholar]
  2. DICKERMAN H. W., REDFIELD B. G., BIERI J. G., WEISSBACH H. STUDIES ON THE ROLE OF VITAMIN B12 FOR THE SYNTHESIS OF METHIONINE IN LIVER. Ann N Y Acad Sci. 1964 Apr 24;112:791–798. doi: 10.1111/j.1749-6632.1964.tb45055.x. [DOI] [PubMed] [Google Scholar]
  3. DICKERMAN H., REDFIELD B. G., BIERI J. G., WEISSBACH H. THE ROLE OF VITAMIN B12 IN METHIONINE BIOSYNTHESIS IN AVIAN LIVER. J Biol Chem. 1964 Aug;239:2545–2552. [PubMed] [Google Scholar]
  4. EAGLE H. Nutrition needs of mammalian cells in tissue culture. Science. 1955 Sep 16;122(3168):501–514. doi: 10.1126/science.122.3168.501. [DOI] [PubMed] [Google Scholar]
  5. Finkelstein J. D., Kyle W., Harris B. J. Methionine metabolism in mammals. Regulation of homocysteine methyltransferases in rat tissue. Arch Biochem Biophys. 1971 Sep;146(1):84–92. doi: 10.1016/s0003-9861(71)80044-9. [DOI] [PubMed] [Google Scholar]
  6. Finkelstein J. D., Mudd S. H. Trans-sulfuration in mammals. The methionine-sparing effect of cystine. J Biol Chem. 1967 Mar 10;242(5):873–880. [PubMed] [Google Scholar]
  7. Goulian M., Beck W. S. Modifications in the Lactobacillus casei assay of serum folate activity. Am J Clin Pathol. 1966 Sep;46(3):390–391. doi: 10.1093/ajcp/46.3_ts.390. [DOI] [PubMed] [Google Scholar]
  8. Kerwar S. S., Spears C., McAuslan B., Weissbach H. Studies on vitamin B12 metabolism in HeLa cells. Arch Biochem Biophys. 1971 Jan;142(1):231–237. doi: 10.1016/0003-9861(71)90279-7. [DOI] [PubMed] [Google Scholar]
  9. Kutzbach C., Galloway E., Stokstad E. L. Influence of vitamin B12 and methionine on levels of folic acid compounds and folate enzymes in rat liver. Proc Soc Exp Biol Med. 1967 Mar;124(3):801–805. doi: 10.3181/00379727-124-31857. [DOI] [PubMed] [Google Scholar]
  10. LOUGHLIN R. E., ELFORD H. L., BUCHANAN J. M. ENZYMATIC SYNTHESIS OF THE METHYL GROUP OF METHIONINE. VII. ISOLATION OF A COBALAMIN-CONTAINING TRANSMETHYLASE (5-METHYLTETRAHYDRO-FOLATE-HOMOCYSTEINE) FROM MAMMALIAN LIVER. J Biol Chem. 1964 Sep;239:2888–2895. [PubMed] [Google Scholar]
  11. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  12. Mangum J. H., Murray B. K., North J. A. Vitamin B 12 dependent methionine biosynthesis in cultured mammalian cells. Biochemistry. 1969 Sep;8(9):3496–3499. doi: 10.1021/bi00837a002. [DOI] [PubMed] [Google Scholar]
  13. Mangum J. H., North J. A. Vitamin B 12-dependent methionine biosynthesis in HEp-2 cells. Biochem Biophys Res Commun. 1968 Jul 11;32(1):105–110. doi: 10.1016/0006-291x(68)90433-6. [DOI] [PubMed] [Google Scholar]
  14. Mudd S. H., Uhlendorf B. W., Hinds K. R. Deranged B 12 metabolism: studies of fibroblasts grown in tissue culture. Biochem Med. 1970 Nov;4(3):215–239. doi: 10.1016/0006-2944(70)90050-5. [DOI] [PubMed] [Google Scholar]
  15. SCHIMKE R. T. ENZYMES OF ARGININE METABOLISM IN MAMMALIAN CELL CULTURE. I. REPRESSION OF ARGININOSUCCINATE SYNTHETASE AND ARGININOSUCCINASE. J Biol Chem. 1964 Jan;239:136–145. [PubMed] [Google Scholar]
  16. Schimke R. T., Doyle D. Control of enzyme levels in animal tissues. Annu Rev Biochem. 1970;39:929–976. doi: 10.1146/annurev.bi.39.070170.004433. [DOI] [PubMed] [Google Scholar]
  17. Vitale J. J., Hegsted D. M. Effects of dietary methionine and vitamin B12 deficiency on folate metabolism. Br J Haematol. 1969 Nov;17(5):467–475. doi: 10.1111/j.1365-2141.1969.tb01395.x. [DOI] [PubMed] [Google Scholar]
  18. WEISSBACH H., PETERKOFSKY A., REDFIELD B. G., DICKERMAN H. STUDIES ON THE TERMINAL REACTION IN THE BIOSYNTHESIS OF METHIONINE. J Biol Chem. 1963 Oct;238:3318–3324. [PubMed] [Google Scholar]
  19. Weissbach H., Taylor R. T. Roles of vitamin B 12 and folic acid in methionine synthesis. Vitam Horm. 1970;28:415–440. doi: 10.1016/s0083-6729(08)60905-x. [DOI] [PubMed] [Google Scholar]

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