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Historical Perspectives: Role of Creatine Phosphate in Energy Transport in Muscle Cells Evaluated by 31 P-NMR. Kazuo Yoshizaki Department of Physiology,

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Presentation on theme: "Historical Perspectives: Role of Creatine Phosphate in Energy Transport in Muscle Cells Evaluated by 31 P-NMR. Kazuo Yoshizaki Department of Physiology,"— Presentation transcript:

1 Historical Perspectives: Role of Creatine Phosphate in Energy Transport in Muscle Cells Evaluated by 31 P-NMR. Kazuo Yoshizaki Department of Physiology, School of Medicine, University of Tokushima, Tokushima , JAPAN

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3 P/O = 2 ~ 3

4 a) ATP carrier model energy-consumption system (actomyosin) energy-production system (mitochondria) energy ATP ADP Pi energy ATP ADP Pi + b ) C~P carrier model Shuttle hypothesis energy-consumption system energy-production system energy ATP ADP Pi C C~P energy ATP ADP C C~P Pi

5 Intracellular Energy Transport in Muscle: Yoshizaki et al BBA 1051:144 (1990) 1) Diffusivities of phosphorus metabolites measured by 31 P-pulsed gradient NMR. 2) The flux of creatine kinase reaction measured by 31 P-saturation transfer NMR. 3) Evaluation on creatine phosphate shuttle theory.

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8 Diffusion coefficients of phosphorus metabolites in frog muscle (D) and in the model solution (D 0 ). D D 0 D/D 0 D/D ATP (x10 -6 cm 2 ·s -1 ) ATP 2.0 ±0.4 (11) 4.5 ±0.6 (7) PCr 2.8 ±0.3 (11) 5.9 ±0.6 (10) Pi 3.3 ±0.3 (11) 7.8 ±1.0 (10) H 2 O 13.6 ±1.0 (5) 22.5* *Mills, R. (1973) J. Phys. Chem

9 a) ATP carrier model energy-consumption system (actomyosin) energy-production system (mitochondria) energy ATP ADP Pi energy ATP ADP Pi + b ) C~P carrier model Shuttle hypothesis energy-consumption system energy-production system energy ATP ADP Pi C C~P energy ATP ADP C C~P Pi

10 Intracellular Energy Transport in Muscle: 1) Diffusivities of phosphorus metabolites measured by 31 P-pulsed gradient NMR. 2) The flux of Creatine kinase reaction measured by 31 P-saturation transfer NMR. 3) Evaluation on creatine phosphate shuttle theory.

11 Dr. G.K. Radda ’ s group at Oxford, U.K., 1983.

12 NMR Spectrometer at Oxford, U.K., 1983

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14 Results of saturation transfer experiments on resting biceps muscle of bullfrog. PCrM + /M 0 T 1m T1T1 kFlux (mmol ・ kg -1 ) (s) (s -1 ) (mmol ・ kg -1 ・ s -1 ) ±2.8±0.03±0.3±0.5±0.03±0.8 The valuse are means and S.D. of 5 muscles.

15 Intracellular Energy Transport in Muscle: 1) Diffusivities of phosphorus metabolites measured by 31 P-pulsed gradient NMR. 2) The flux of Creatine kinase reaction measured by 31 P-saturation transfer NMR. 3) Evaluation on creatine phosphate shuttle theory.

16 Diffusion distance τ: life time D: diffusion coefficient 2Dτ

17 CPK flux PCr + ADP Cr + ATP Estimation of life times of substrates for creatine kinase reaction in resting biceps muscle of bullfrog. Substrates PCr ADP Cr ATP Concentrations x (mmol·kg -1 ) Life times (s) x Life time = Concentration / the flux of the creatine kinase reaction (3.6 mmol·kg -1 ·s -1 ).

18 Diffusion distance (μm) 22 μm Dτ

19 Newsholme and Start (1973)

20 mitochondriamyofibril ADP ATP

21 A case report on acute myocardial infarction accompanied with no increase in serum creatine kinase activity. K. Koizumi et al., Kobe City General Hospital, J. Jap. Coll. Angiol., 27, 988 (1987) (in Japanese) A 56 years old female, suffering from Diabetes mellitus without cardiac complications, died with acute myocardiac infraction. The agar gel electrophoresis of muscle homogenates showed that the CK-MM isozyme was almost undetected in skeletal and cardiac muscles, and the CK-MB isozyme was decreased in cardiac muscle. (Tentatively translated by K. Yoshizaki) P 2-4 血清CK上昇を伴わない急性心筋梗塞症の一例 神戸市立中央市民病院 循環器センター 同臨床病理科 * 小泉克己, 吉川純一, 加藤 洋, 奥町冨久丸, 吉田 清, 白鳥健一, 高尾精一, 赤阪隆史, 赤土正洋, 城 泰子, 笠倉新平 * , 内田博也 *

22 a) ATP carrier model energy-consumption system (actomyosin) energy-production system (mitochondria) energy ATP ADP Pi energy ATP ADP Pi + b ) C~P carrier model Shuttle hypothesis energy-consumption system energy-production system energy ATP ADP Pi C C~P energy ATP ADP C C~P Pi

23 In vivo regulation of mitochondrial respiration in cardiomyocytes: Specific restrictions for intracellular diffusion of ADP Valdur A. Sakes, Yulia O. Belikova and Andrey V. Kuznetsov Laboratory of Bioenergetics, U. S. S. R. Cardiology Research Center, Moscow (U. S. S. R.) Biochmica et Biophysica Acta, 1074 (1991) Preparations K m (μM) K m (mM) ADP creatine no additions +AP 5 A +(creatine) (at 0.2mM ATP) Mitochondria 17.6± ± ±0.14 Saponin-treated carduinyocytes 250.2± ± ± ±0.11 Skinned fibers 263.7± ±8 6.25±0.21

24 In vivo regulation of mitochondrial respiration in cardiomyocytes: Specific restrictions for intracellular diffusion of ADP Valdur A. Sakes, Yulia O. Belikova and Andrey V. Kuznetsov Laboratory of Bioenergetics, U. S. S. R. Cardiology Research Center, Moscow (U. S. S. R.) Biochmica et Biophysica Acta, 1074 (1991) Preparations K m (μM) K m (mM) ADP creatine no additions +AP 5 A +(creatine) (at 0.2mM ATP) Mitochondria 17.6± ± ±0.14 Saponin-treated carduinyocytes 250.2± ± ± ±0.11 Skinned fibers 263.7± ±8 6.25±0.21 Discussion: (The old Japanese lady, described by Oita et al. (1988) reported to have only 2% of intact creatine kinase preserved, might not be able to perform too much of work any more and might had suffered from decreased relaxation; however, such a case should be verified by more numerous studies to be taken seriously.)

25 Nicolay et al. (1998) Mol. Cell. Biochem 184: P-saturation transfer NMR on skeletal muscles of creatine kinase (CK)-knockout mice. wild M-CK(-/-)

26 Nicolay et al. (1998) Mol. Cell. Biochem 184: P-saturation transfer NMR on skeletal muscles of creatine kinase (CK)-knockout mice. wild M-CK(-/-)

27 1983

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