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. 1982 Mar 15;202(3):573–579. doi: 10.1042/bj2020573

A p.m.r. isotope-exchange method for studying the kinetic properties of dehydrogenases in intact cells.

R J Simpson, K M Brindle, F F Brown, I D Campbell, D L Foxall
PMCID: PMC1158150  PMID: 7092831

Abstract

A method to determine the activity of dehydrogenases in an intact-cell system is described. The method involves the use of n.m.r. to monitor bulk isotope exchange. The approach is illustrated by application to the isotope equilibration of pyruvate and lactate as catalyzed by lactate dehydrogenase in intact erythrocytes. Particular problems peculiar to bulk isotope exchange and its observation by n.m.r. are considered.

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Selected References

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

  1. Borgmann U., Moon T. W., Laidler K. J. Molecular kinetics of beef heart lactate dehydrogenase. Biochemistry. 1974 Dec 3;13(25):5152–5158. doi: 10.1021/bi00722a016. [DOI] [PubMed] [Google Scholar]
  2. Brindle K. M., Brown F. F., Campbell I. D., Foxall D. L., Simpson R. J. 1H/2H isotope exchange studies in intact erythrocytes. Biochem Soc Trans. 1980 Oct;8(5):646–647. doi: 10.1042/bst0080646. [DOI] [PubMed] [Google Scholar]
  3. Brindle K. M., Brown F. F., Campbell I. D., Grathwohl C., Kuchel P. W. Application of spin-echo nuclear magnetic resonance to whole-cell systems. Membrane transport. Biochem J. 1979 Apr 15;180(1):37–44. doi: 10.1042/bj1800037. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Brown F. F., Campbell I. D., Kuchel P. W., Rabenstein D. C. Human erythrocyte metabolism studies by 1H spin echo NMR. FEBS Lett. 1977 Oct 1;82(1):12–16. doi: 10.1016/0014-5793(77)80875-2. [DOI] [PubMed] [Google Scholar]
  5. Campbell I. D., Dobson C. M. The application of high resolution nuclear magnetic resonance to biological systems. Methods Biochem Anal. 1979;25:1–133. doi: 10.1002/9780470110454.ch1. [DOI] [PubMed] [Google Scholar]
  6. Clark B. R. Fluorometric quantitation of picomole amounts of 1-methylnicotinamide and nicotinamide in serum. Methods Enzymol. 1980;66:5–8. doi: 10.1016/0076-6879(80)66432-5. [DOI] [PubMed] [Google Scholar]
  7. Cook P. F., Oppenheimer N. J., Cleland W. W. Secondary deuterium and nitrogen-15 isotope effects in enzyme-catalyzed reactions. Chemical mechanism of liver alcohol dehydrogenase. Biochemistry. 1981 Mar 31;20(7):1817–1825. doi: 10.1021/bi00510a016. [DOI] [PubMed] [Google Scholar]
  8. Deuticke B., Rickert I., Beyer E. Stereoselective, SH-dependent transfer of lactate in mammalian erythrocytes. Biochim Biophys Acta. 1978 Feb 2;507(1):137–155. doi: 10.1016/0005-2736(78)90381-4. [DOI] [PubMed] [Google Scholar]
  9. Everse J., Kaplan N. O. Lactate dehydrogenases: structure and function. Adv Enzymol Relat Areas Mol Biol. 1973;37:61–133. doi: 10.1002/9780470122822.ch2. [DOI] [PubMed] [Google Scholar]
  10. Halestrap A. P. Transport of pyruvate nad lactate into human erythrocytes. Evidence for the involvement of the chloride carrier and a chloride-independent carrier. Biochem J. 1976 May 15;156(2):193–207. doi: 10.1042/bj1560193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Marshall W. E., Omachi A. Measured and calculated NAD+-NADH ratios in human erythrocytes. Biochim Biophys Acta. 1974 Jun 20;354(1):1–10. doi: 10.1016/0304-4165(74)90046-4. [DOI] [PubMed] [Google Scholar]
  12. Rivedal E., Sanner T. Role of ions in the regulation of porcine lactate dehydrogenase. Biochim Biophys Acta. 1979 Mar 16;567(1):60–65. doi: 10.1016/0005-2744(79)90172-4. [DOI] [PubMed] [Google Scholar]
  13. SILVERSTEIN E., BOYER P. D. EQUILIBRIUM REACTION RATES AND THE MECHANISMS OF BOVINE HEART AND RABBIT MUSCLE LACTATE DEHYDROGENASES. J Biol Chem. 1964 Nov;239:3901–3907. [PubMed] [Google Scholar]
  14. Simpson R. J., Brindle K. M., Brown F. F., Campbell I. D., Foxall D. L. Studies of lactate dehydrogenase in the purified state and in intact erythrocytes. Biochem J. 1982 Mar 15;202(3):581–587. doi: 10.1042/bj2020581. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Simpson R. J., Brindle K. M., Brown F. F., Campbell I. D., Foxall D. L. Studies of pyruvate-water isotope exchange catalysed by erythrocytes and proteins. Biochem J. 1981 Feb 1;193(2):401–406. doi: 10.1042/bj1930401. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Trommer W. E., Huth H., Wenzel H. R. The nature of the substrate inhibition in lactate dehydrogenases as studied by a spin-labeled derivative of NAD. Biochim Biophys Acta. 1979 Mar 16;567(1):49–59. doi: 10.1016/0005-2744(79)90171-2. [DOI] [PubMed] [Google Scholar]
  17. VONKORFF R. W. PYRUVATE-C14, PURITY AND STABILITY. Anal Biochem. 1964 Jun;8:171–178. doi: 10.1016/0003-2697(64)90043-0. [DOI] [PubMed] [Google Scholar]
  18. WHITTAM R. Potassium movements and ATP in human red cells. J Physiol. 1958 Mar 11;140(3):479–497. [PMC free article] [PubMed] [Google Scholar]
  19. Williamson D. H., Lund P., Krebs H. A. The redox state of free nicotinamide-adenine dinucleotide in the cytoplasm and mitochondria of rat liver. Biochem J. 1967 May;103(2):514–527. doi: 10.1042/bj1030514. [DOI] [PMC free article] [PubMed] [Google Scholar]

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