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. 1985 Aug;365:259–276. doi: 10.1113/jphysiol.1985.sp015770

Dependence of adenosine triphosphatase activity of rabbit psoas muscle fibres and myofibrils on substrate concentration.

H Glyn, J Sleep
PMCID: PMC1193000  PMID: 3162018

Abstract

The rate of hydrolysis of adenosine triphosphate (ATP) by chemically skinned rabbit muscle fibres was measured as a function of Mg ATP concentration in the range 5 microM to 5 mM. Pyruvate kinase and lactate dehydrogenase were used to link adenosine diphosphate formation to oxidation of nicotinamide adenine dinucleotide which was followed by the change in absorption at 340 nm. The ATPase rate of a fully activated fibre (pCa = 4.5) increased monotonically with Mg ATP concentration in a manner that could be readily fitted by a hyperbola. At 15 degrees C, pH 7 and an ionic strength of 0.2 M the rate at saturating Mg ATP (Vm) was 1.78 +/- 0.2 s-1 per myosin head (mean +/- S.D.; n = 6) and the Mg ATP concentration needed for half the maximal rate (Km) was 16.6 +/- 2 microM. The ATPase of fibres that had been stabilized by cross-linking with 1-ethyl-3-(3-dimethyl-aminopropyl)carbodiimide (EDC) was also investigated. Cross-linking did not significantly affect the Vm or Km and these fibres proved useful for investigating the adequacy of the pyruvate kinase activity for regenerating hydrolysed ATP. Myofibrils were cross-linked with EDC or glutaraldehyde to prevent shortening. Their ATPase properties were investigated: the values of Vm were 0.85 +/- 0.18 (mean +/- S.D.; n = 14) and 0.82 +/- 0.05 s-1 (n = 6) and of Km were 18.0 +/- 2.8 and 12.4 +/- 2.4 microM respectively. The values of Vm and Km for EDC cross-linked myofibrils were fairly insensitive to ionic strength, the Km decreasing 40% and the Vm increasing 50% for a change from 0.2 to 0.3 M. This slight dependence on ionic strength is considered in relation to the ionic strength dependence of the elementary rate constants of the actomyosin subfragment-1 ATPase cycle.

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Selected References

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  1. Brenner B., Schoenberg M., Chalovich J. M., Greene L. E., Eisenberg E. Evidence for cross-bridge attachment in relaxed muscle at low ionic strength. Proc Natl Acad Sci U S A. 1982 Dec;79(23):7288–7291. doi: 10.1073/pnas.79.23.7288. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Cooke R., Bialek W. Contraction of glycerinated muscle fibers as a function of the ATP concentration. Biophys J. 1979 Nov;28(2):241–258. doi: 10.1016/S0006-3495(79)85174-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Eisenberg E., Hill T. L., Chen Y. Cross-bridge model of muscle contraction. Quantitative analysis. Biophys J. 1980 Feb;29(2):195–227. doi: 10.1016/S0006-3495(80)85126-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Eisenberg E., Moos C. Actin activation of heavy meromyosin adenosine triphosphatase. Dependence on adenosine triphosphate and actin concentrations. J Biol Chem. 1970 May 10;245(9):2451–2456. [PubMed] [Google Scholar]
  5. Ferenczi M. A., Goldman Y. E., Simmons R. M. The dependence of force and shortening velocity on substrate concentration in skinned muscle fibres from Rana temporaria. J Physiol. 1984 May;350:519–543. doi: 10.1113/jphysiol.1984.sp015216. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Ferenczi M. A., Homsher E., Trentham D. R. The kinetics of magnesium adenosine triphosphate cleavage in skinned muscle fibres of the rabbit. J Physiol. 1984 Jul;352:575–599. doi: 10.1113/jphysiol.1984.sp015311. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Gillis J. M., Maréchal G. The incorporation of radioactive phosphate into ATP in glycerinated fibres stretched or released during contraction. J Mechanochem Cell Motil. 1974;3(1):55–68. [PubMed] [Google Scholar]
  8. Goldman Y. E., Hibberd M. G., Trentham D. R. Initiation of active contraction by photogeneration of adenosine-5'-triphosphate in rabbit psoas muscle fibres. J Physiol. 1984 Sep;354:605–624. doi: 10.1113/jphysiol.1984.sp015395. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Goodno C. C., Wall C. M., Perry S. V. Kinetics and regulation of the myofibrillar adenosine triphosphatase. Biochem J. 1978 Dec 1;175(3):813–821. doi: 10.1042/bj1750813. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Knight P. J., Trinick J. A. Preparation of myofibrils. Methods Enzymol. 1982;85(Pt B):9–12. doi: 10.1016/0076-6879(82)85004-0. [DOI] [PubMed] [Google Scholar]
  11. Marston S. B., Tregear R. T. Nucleotide binding to myosin in calcium activated muscle. Biochim Biophys Acta. 1974 Mar 26;333(3):581–584. doi: 10.1016/0005-2728(74)90143-1. [DOI] [PubMed] [Google Scholar]
  12. Maruyama K., Weber A. Binding of adenosine triphosphate to myofibrils during contraction and relaxation. Biochemistry. 1972 Aug 1;11(16):2990–2998. doi: 10.1021/bi00766a010. [DOI] [PubMed] [Google Scholar]
  13. Mornet D., Bertrand R., Pantel P., Audemard E., Kassab R. Structure of the actin-myosin interface. Nature. 1981 Jul 23;292(5821):301–306. doi: 10.1038/292301a0. [DOI] [PubMed] [Google Scholar]
  14. Scopes R. K. Purification of glycolytic enzymes by using affinity-elution chromatography. Biochem J. 1977 Feb 1;161(2):253–263. doi: 10.1042/bj1610253. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Sleep J. A. Single turnovers of adenosine 5'-triphosphate by myofibrils and actomyosin subfragment 1. Biochemistry. 1981 Aug 18;20(17):5043–5051. doi: 10.1021/bi00520a034. [DOI] [PubMed] [Google Scholar]
  16. Stein L. A., Chock P. B., Eisenberg E. Mechanism of the actomyosin ATPase: effect of actin on the ATP hydrolysis step. Proc Natl Acad Sci U S A. 1981 Mar;78(3):1346–1350. doi: 10.1073/pnas.78.3.1346. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Takashi R., Putnam S. A fluorimetric method for continuously assaying ATPase: application to small specimens of glycerol-extracted muscle fibers. Anal Biochem. 1979 Jan 15;92(2):375–382. doi: 10.1016/0003-2697(79)90674-2. [DOI] [PubMed] [Google Scholar]
  18. Taylor E. W. Transient phase of adenosine triphosphate hydrolysis by myosin, heavy meromyosin, and subfragment 1. Biochemistry. 1977 Feb 22;16(4):732–739. doi: 10.1021/bi00623a027. [DOI] [PubMed] [Google Scholar]
  19. Ulbrich M., Rüegg J. C. Is the chemomechanical energy transformation reversible? Pflugers Arch. 1976 Jun 22;363(3):219–222. doi: 10.1007/BF00594604. [DOI] [PubMed] [Google Scholar]
  20. Weber A., Murray J. M. Molecular control mechanisms in muscle contraction. Physiol Rev. 1973 Jul;53(3):612–673. doi: 10.1152/physrev.1973.53.3.612. [DOI] [PubMed] [Google Scholar]
  21. White H. D., Taylor E. W. Energetics and mechanism of actomyosin adenosine triphosphatase. Biochemistry. 1976 Dec 28;15(26):5818–5826. doi: 10.1021/bi00671a020. [DOI] [PubMed] [Google Scholar]
  22. Wood D. S., Zollman J., Reuben J. P., Brandt P. W. Human skeletal muscle: properties of the "chemically skinned%" fiber. Science. 1975 Mar 21;187(4181):1075–1076. doi: 10.1126/science.187.4181.1075. [DOI] [PubMed] [Google Scholar]
  23. Yates L. D., Greaser M. L. Quantitative determination of myosin and actin in rabbit skeletal muscle. J Mol Biol. 1983 Jul 25;168(1):123–141. doi: 10.1016/s0022-2836(83)80326-x. [DOI] [PubMed] [Google Scholar]

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