Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1975 Feb;121(2):640–647. doi: 10.1128/jb.121.2.640-647.1975

Enzymes of deoxythymidine triphosphate biosynthesis in Neurospora crassa mitochondria.

M Rossi, D O Woodward
PMCID: PMC245976  PMID: 163227

Abstract

Intact mitochondria of Neurospora crassa incorporate deoxythymidine 5'-monophosphate (dTMP) into deoxyribonucleic acid but not the label from (methyl-3H) deoxythymidine. Mitochondrial homogenates contain deoxythymidylate kinase (EC 2.7.4.9), deoxycytidylate aminohydrolase (dCMP deaminase) (EC 3.5.4.12), and thymidylate synthetase (EC 2.1.1b), but not thymidine kinase (EC 2.7.1.21) activity. dTMP kinase is loosely bound to the mitochondrial membrane and is solubilized by 0.4 M KCl in mitochondrial homogenates, the dCMP aminohydrolase deaminase) is bound to the inner membrane and is not solubilized by 0.4 M KCl. dTMP synthetase activity is found in the 2,000 times g particulate fractions by homogenization of mitochondria in 0.4 M KCl. The dCMP deaminase activity found in the particulate fraction of the inner membrane is efficiently regulated by the products of the pathway: deoxycytidine 5'-triphosphate activates whereas deoxythymidine 5'-triphosphate inhibits, as found for the soluble enzyme from other sources. These data indicate that mitochondria of N. crassa contain specific enzymes for the biosynthesis of deoxythymidine triphosphate.

Full text

PDF
642

Selected References

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

  1. Ashwell M., Work T. S. The biogenesis of mitochondria. Annu Rev Biochem. 1970;39:251–290. doi: 10.1146/annurev.bi.39.070170.001343. [DOI] [PubMed] [Google Scholar]
  2. BURTON K. A study of the conditions and mechanism of the diphenylamine reaction for the colorimetric estimation of deoxyribonucleic acid. Biochem J. 1956 Feb;62(2):315–323. doi: 10.1042/bj0620315. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Berk A. J., Clayton D. A. A genetically distinct thymidine kinase in mammalian mitochondria. Exclusive labeling of mitochondrial deoxyribonucleic acid. J Biol Chem. 1973 Apr 25;248(8):2722–2729. [PubMed] [Google Scholar]
  4. Borst P., Kroon A. M. Mitochondrial DNA: physicochemical properties, replication, and genetic function. Int Rev Cytol. 1969;26:107–190. doi: 10.1016/s0074-7696(08)61635-6. [DOI] [PubMed] [Google Scholar]
  5. Dunlap R. B., Harding N. G., Huennekens F. M. Thymidylate synthetase from amethopterin-resistant Lactobacillus casei. Biochemistry. 1971 Jan 5;10(1):88–97. doi: 10.1021/bi00777a014. [DOI] [PubMed] [Google Scholar]
  6. FINK R. M., FINK K. Relative retention of H3 and C14 labels of nucleosides incorporated into nucleic acids of Neurospora. J Biol Chem. 1962 Sep;237:2889–2891. [PubMed] [Google Scholar]
  7. Granick S., Gibor A. The DNA of chloroplasts, mitochondria and centrioles. Prog Nucleic Acid Res Mol Biol. 1967;6:143–186. doi: 10.1016/s0079-6603(08)60526-7. [DOI] [PubMed] [Google Scholar]
  8. Grivell A. R., Jackson J. F. Thymidine kinase: evidence for its absence from Neurospora crassa and some other micro-organisms, and the relevance of this to the specific labelling of deoxyribonucleic acid. J Gen Microbiol. 1968 Dec;54(2):307–317. doi: 10.1099/00221287-54-2-307. [DOI] [PubMed] [Google Scholar]
  9. Iwashima A., Rabinowitz M. Partial purification of mitochondrial and supernatant DNA polymerase from Saccharomyces cerevisiae. Biochim Biophys Acta. 1969 Apr 22;178(2):283–293. doi: 10.1016/0005-2744(69)90396-9. [DOI] [PubMed] [Google Scholar]
  10. KIELLEY R. K. Particle-bound thymidylate kinase in mouse liver, a possible factor in the control of DNA synthesis. Biochem Biophys Res Commun. 1963 Feb 6;10:249–253. doi: 10.1016/0006-291x(63)90425-x. [DOI] [PubMed] [Google Scholar]
  11. Kalf G. F., Ch'ih J. J. Purification and properties of deoxyribonucleic acid polymerase from rat liver mitochondria. J Biol Chem. 1968 Sep 25;243(18):4904–4916. [PubMed] [Google Scholar]
  12. Küntzel H., Schäfer K. P. Mitochondrial RNA polymerase from Neurospora crassa. Nat New Biol. 1971 Jun 30;231(26):265–269. doi: 10.1038/newbio231265a0. [DOI] [PubMed] [Google Scholar]
  13. 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]
  14. LUCK D. J., REICH E. DNA IN MITOCHONDRIA OF NEUROSPORA CRASSA. Proc Natl Acad Sci U S A. 1964 Oct;52:931–938. doi: 10.1073/pnas.52.4.931. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Larsson A., Reichard P. Enzymatic synthesis of deoxyribonucleotides. IX. Allosteric effects in the reduction of pyrimidine ribonucleotides by the ribonucleoside diphosphate reductase system of Escherichia coli. J Biol Chem. 1966 Jun 10;241(11):2533–2539. [PubMed] [Google Scholar]
  16. Mitra R. S., Bernstein I. A. Thymidine incorporation into deoxyribonucleic acid by isolated rat liver mitochondria. J Biol Chem. 1970 Mar 25;245(6):1255–1260. [PubMed] [Google Scholar]
  17. Munro H. N. The determination of nucleic acids. Methods Biochem Anal. 1966;14:113–176. doi: 10.1002/9780470110324.ch5. [DOI] [PubMed] [Google Scholar]
  18. O'Donovan G. A., Neuhard J. Pyrimidine metabolism in microorganisms. Bacteriol Rev. 1970 Sep;34(3):278–343. doi: 10.1128/br.34.3.278-343.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. OKAZAKI R., KORNBERG A. DEOXYTHYMIDINE KINASE OF ESCHERICHIA COLI. II. KINETICS AND FEEDBACK CONTROL. J Biol Chem. 1964 Jan;239:275–284. [PubMed] [Google Scholar]
  20. Parsons P., Simpson M. V. Biosynthesis of DNA by isolated mitochondria: incorporation of thymidine triphosphate-2-C-14. Science. 1967 Jan 6;155(3758):91–93. doi: 10.1126/science.155.3758.91. [DOI] [PubMed] [Google Scholar]
  21. Rossi M., Geraci G., Scarano E. Deoxycytidylate aminohydrolase. 3. Modifications of the substrate sites caused by allosteric effectors. Biochemistry. 1967 Dec;6(12):3640–3645. doi: 10.1021/bi00864a003. [DOI] [PubMed] [Google Scholar]
  22. Schnaitman C., Greenawalt J. W. Enzymatic properties of the inner and outer membranes of rat liver mitochondria. J Cell Biol. 1968 Jul;38(1):158–175. doi: 10.1083/jcb.38.1.158. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Sergott R. C., Debeer L. J., Bessman M. J. On the regulation of a bacterial deoxycytidylate deaminase. J Biol Chem. 1971 Dec 25;246(24):7755–7758. [PubMed] [Google Scholar]
  24. Suyama Y., Eyer J. Ribonucleic acid synthesis in isolated mitochondria from Tetrahymena. J Biol Chem. 1968 Jan 25;243(2):320–328. [PubMed] [Google Scholar]
  25. Tsai M. J., Michaelis G., Criddle R. S. DNA-dependent RNA polymerase from yeast mitochondria. Proc Natl Acad Sci U S A. 1971 Feb;68(2):473–477. doi: 10.1073/pnas.68.2.473. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. WINTERSBERGER E. DNA-ABHAENGIGE RNA-SYNTHESE IN RATTENLEBER-MITOCHONDRIEN. Hoppe Seylers Z Physiol Chem. 1964;336:285–288. doi: 10.1515/bchm2.1964.336.1.285. [DOI] [PubMed] [Google Scholar]
  27. Zollinger W. D., Woodward D. O. Comparison of cysteine and tryptophan content of insoluble proteins derived from wild-type and mi-1 strains of Neurospora crassa. J Bacteriol. 1972 Mar;109(3):1001–1013. doi: 10.1128/jb.109.3.1001-1013.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES