Folyl and Antifolyl Polyglutamates pp 183-197 | Cite as
Folylpolyglutamate Synthetase Activities of Neurospora Crassa: Nature of Products Formed by Soluble and Particulate Enzymes in the Wild Type and Polyglutamate-Deficient Mutants
Abstract
The folylpolyglutamate synthetase activities of Neurospora crassa wild type (FGSC 853) and two polyglutamate-deficient mutants (met-6, 35809, FGSC 1330 and mac, 65108, FGSC 3609) were examined using dialyzed extracts prepared during exponential mycelial growth. Enzyme assay was based on incorporation of [U-3H]glutamate in folyl-polyglutamates that were separated by gradient elution from DEAE-cellulose. Extracts of the wild type produced H4PteGlu2 (15%), H4PteGlu3 (35%) and H4PteGlu6 (50%) when anaerobically incubated with glutamate, ATP, and H4PteGlu. Under these conditions, the met-6 produced only H4PteGlu2 and higher polyglutamates (H4PteGlu4 and H4PteGlu5) were not utilized. The mac mutant failed to catalyze addition of glutamate to H4PteGlu. However, H4PteGlu2 was effectively converted to the tri-, and hexaglutamates. Mixing wild type and met-6 protein stimulated the formation of tri-, and hexaglutamates. Mixing mac and met-6 extracts resulted in H4PteGlua and H4PteGlu6 labeling when glutamate and H4PteGlu were provided.
Keywords
Potassium Phosphate Wild Type Protein Neurospora CRASSA Synthetase Activity 2S04 FractionPreview
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References
- 1.Gawthorne, J. M., and Smith, R. M., Biochem. J., 136, 295–301 (1973).PubMedGoogle Scholar
- 2.Taylor, R. T., and Hanna, M. L., Arch. Biochem. Biophys., 181, 331–344 (1977).PubMedCrossRefGoogle Scholar
- 3.Taylor, R. T., and Hanna, M. L., Arch. Biochem. Biophys., 197, 36–43 (1979).PubMedCrossRefGoogle Scholar
- 4.McGuire, J. J., Kitamoto, Y., Hsieh, P., Coward, J. K., and Bertino, J. R., in: Chemistry and Biology of Pteridines (R. L. Kisliuk and G. M. Brown, eds.), pp. 471–476, Elsevier-North Holland, New York (1979).Google Scholar
- 5.Masurekar, M., and Brown, G. M., Biochemistry, 14, 2424–2430 (1975).PubMedCrossRefGoogle Scholar
- 6.Shane, B., Brody, T., and Stokstad, E. L. R., in: Chemistry and Biology of Pteridines (R. L. Kisliuk, and G. M. Brown, eds.), pp. 341–346, Elsevier-North Holland, New York (1979).Google Scholar
- 7.Shane, B., J. Biol. Chem., 255, 5655–5662 (1980).PubMedGoogle Scholar
- 8.Shane, B., J. Biol. Chem., 255, 5663–5667 (1980).PubMedGoogle Scholar
- 9.Schetel, M. F., Boehue, J. W., and Libby, D. A., J. Biol. Chem., 240, 3154–3158 (1965).Google Scholar
- 10.Lor, K. L., and Cossins, E. A., Biochem. J., 130, 773–783 (1972).PubMedGoogle Scholar
- 11.Baugh, C. M., Braverman, E., and Nair, M. G., in: Chemistry and Biology of Pteridines (W. Pfleiderer, ed.), pp. 465–474, Walter de Gruyter, Berlin (1975).Google Scholar
- 12.Bassett, J. R., Weir, D. G., and Scott, J. M., J. Gen. Microbiol., 93, 169–172 (1976).PubMedCrossRefGoogle Scholar
- 13.Cossins, E. A., Chan, P.-Y., and Combepine, G., Biochem. J., 160, 305–314 (1976).PubMedGoogle Scholar
- 14.Cossins, E. A., and Chan, P. Y., Plant Biochemical J. S. M. Sircar Mem. Vol., 53–62 (1980).Google Scholar
- 15.Chan, P. Y., and Cossins, E. A., Arch. Biochem. Biophys., 200, 346–356 (1980).PubMedCrossRefGoogle Scholar
- 16.Sakami, W., Ritari, S. J., Black, C. W., and Rzepka, J., Fed. Proc, 32, 471 (1973).Google Scholar
- 17.Ritari, S. J., Sakami, W., Black, C. W., and Rzepka, J. Anal. Biochem., 63, 118–129 (1975).PubMedCrossRefGoogle Scholar
- 18.Ritari, S. J., Sakami, W., Black, C. W., and Rzepka, J., Neuro-spora Newletter, 20, 26–27 (1973).Google Scholar
- 19.Ritari, S. J., Sakami, W., and Black, C. W., Neurospora New-letter, 20, 27 (1973).Google Scholar
- 20.Reid, V. E., and Friedkin, M., Mol. Pharmacol., 9, 74–80 (1973).PubMedGoogle Scholar
- 21.Vogel, J. H., Amer. Natur., 98, 435–446 (1964).CrossRefGoogle Scholar
- 22.Kuntzel, H., and Noll, H., Nature, 215, 1340–1345 (1967).PubMedCrossRefGoogle Scholar
- 23.Lowry, O. H., Rosebrough, N. J., Farr, A. L., and Randall, R. J., J. Biol. Chem., 193, 265–275 (1951).PubMedGoogle Scholar
- 24.Clandinin, M. T., and Cossins, E. A., Biochem. J., 128, 29–40 (1972).PubMedGoogle Scholar
- 25.Turner, J. C, Int. J. Appl. Radiat. Isotop., 20, 499–505 (1969).CrossRefGoogle Scholar
- 26.Covey, J. M., Life Sciences, 26, 665–678 (1980).PubMedCrossRefGoogle Scholar
- 27.Cossins, E. A., in: The Biochemistry of Plants, Vol. 2, (D. D. Davis, ed.), pp. 365–418, Academic Press, New York (1980).Google Scholar
- 28.Zelikson, R., and Luzzati, M., Eur. J. Biochem., 79, 285–292 (1977).PubMedCrossRefGoogle Scholar
- 29.Murray, N. E., Genetics, 61, 67–77 (1969).PubMedGoogle Scholar
- 30.Burton, E., Selhub, J., and Sakami, W., Biochem. J., 111, 793–795 (1969).PubMedGoogle Scholar