Difference between revisions of "Perry 2011 Biochem J"
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{{Publication | {{Publication | ||
|title=Perry CG, Kane DA, Lin CT, Kozy R, Cathey BL, Lark DS, Kane CL, Brophy PM, Gavin TP, Anderson EJ, Neufer PD (2011) Inhibiting myosin-ATPase reveals a dynamic range of mitochondrial respiratory control in skeletal muscle. Biochem J 437: 215- | |title=Perry CG, Kane DA, Lin CT, Kozy R, Cathey BL, Lark DS, Kane CL, Brophy PM, Gavin TP, Anderson EJ, Neufer PD (2011) Inhibiting myosin-ATPase reveals a dynamic range of mitochondrial respiratory control in skeletal muscle. Biochem J 437:215-22. | ||
|info=[http://www.ncbi.nlm.nih.gov/pubmed/21554250 PMID:21554250] | |info=[http://www.ncbi.nlm.nih.gov/pubmed/21554250 PMID:21554250 Open Access] | ||
|authors=Perry CG, Kane DA, Lin CT, Kozy R, Cathey BL, Lark DS, Kane CL, Brophy PM, Gavin TP, Anderson EJ, Neufer PD | |authors=Perry CG, Kane DA, Lin CT, Kozy R, Cathey BL, Lark DS, Kane CL, Brophy PM, Gavin TP, Anderson EJ, Neufer PD | ||
|year=2011 | |year=2011 | ||
|journal=Biochem J | |journal=Biochem J | ||
|abstract=Assessment of mitochondrial ADP-stimulated respiratory kinetics in | |abstract=Assessment of mitochondrial ADP-stimulated respiratory kinetics in permeabilized fibre (pfi) bundles is increasingly used in clinical diagnostic and basic research settings. However, estimates of the ''K''<sub>m</sub> for ADP vary considerably (~20-300 μM) and tend to overestimate respiration at rest. Noting that pfi bundles spontaneously contract during respiration experiments, we systematically determined the impact of contraction, temperature and oxygenation on ADP-stimulated respiratory kinetics. BLEB ('''blebbistatin'''), a myosin II ATPase inhibitor, blocked contraction under all conditions and yielded high ''K''<sub>m</sub> values for ADP of >~250 and ~80 μM in red and white rat pfi bundles respectively. In the absence of BLEB, pfi bundles contracted and the ''K''<sub>m</sub> for ADP decreased ~2-10-fold in a temperature-dependent manner. pfi bundles were sensitive to hyperoxia (increased ''K''<sub>m</sub>) in the absence of BLEB (contracted) at 30 °C but not 37 °C. In pfi bundles from humans, contraction elicited high sensitivity to ADP (''K''<sub>m</sub><100 μM), whereas blocking contraction (+BLEB) and including a phosphocreatine/creatine ratio of 2:1 to mimic the resting energetic state yielded a ''K''<sub>m</sub> for ADP of ~1560 μM, consistent with estimates of ''in vivo'' resting respiratory rates of <1% maximum. These results demonstrate that the sensitivity of muscle to ADP varies over a wide range in relation to contractile state and cellular energy charge, providing evidence that enzymatic coupling of energy transfer within skeletal muscle becomes more efficient in the working state. | ||
|keywords= | |keywords=Bioenergetics, Blebbistatin, Creatine kinase, Myosin-ATPase, N-benzyltoluene sulfonamide, Skeletal muscle contraction | ||
|mipnetlab=US_NC Greenville_Neufer PD, CA Toronto Perry CG, US_NC_Greenville_Anderson EJ, CA_Antigonish_Kane DA | |mipnetlab=US_NC Greenville_Neufer PD, CA Toronto Perry CG, US_NC_Greenville_Anderson EJ, CA_Antigonish_Kane DA | ||
}} | }} | ||
{{Labeling | {{Labeling | ||
| | |area=Respiration | ||
|organism=Human, Rat | |organism=Human, Rat | ||
|tissues=Skeletal muscle | |tissues=Skeletal muscle | ||
|preparations=Permeabilized tissue | |preparations=Permeabilized tissue | ||
|enzymes=Complex I, Complex V;ATP synthase | |||
|topics=ADP, Inhibitor, Temperature | |||
|pathways=N | |||
|instruments=Oxygraph-2k | |||
|additional=Blebbistatin, | |||
}} | }} |
Latest revision as of 16:12, 6 March 2020
Perry CG, Kane DA, Lin CT, Kozy R, Cathey BL, Lark DS, Kane CL, Brophy PM, Gavin TP, Anderson EJ, Neufer PD (2011) Inhibiting myosin-ATPase reveals a dynamic range of mitochondrial respiratory control in skeletal muscle. Biochem J 437:215-22. |
Perry CG, Kane DA, Lin CT, Kozy R, Cathey BL, Lark DS, Kane CL, Brophy PM, Gavin TP, Anderson EJ, Neufer PD (2011) Biochem J
Abstract: Assessment of mitochondrial ADP-stimulated respiratory kinetics in permeabilized fibre (pfi) bundles is increasingly used in clinical diagnostic and basic research settings. However, estimates of the Km for ADP vary considerably (~20-300 μM) and tend to overestimate respiration at rest. Noting that pfi bundles spontaneously contract during respiration experiments, we systematically determined the impact of contraction, temperature and oxygenation on ADP-stimulated respiratory kinetics. BLEB (blebbistatin), a myosin II ATPase inhibitor, blocked contraction under all conditions and yielded high Km values for ADP of >~250 and ~80 μM in red and white rat pfi bundles respectively. In the absence of BLEB, pfi bundles contracted and the Km for ADP decreased ~2-10-fold in a temperature-dependent manner. pfi bundles were sensitive to hyperoxia (increased Km) in the absence of BLEB (contracted) at 30 °C but not 37 °C. In pfi bundles from humans, contraction elicited high sensitivity to ADP (Km<100 μM), whereas blocking contraction (+BLEB) and including a phosphocreatine/creatine ratio of 2:1 to mimic the resting energetic state yielded a Km for ADP of ~1560 μM, consistent with estimates of in vivo resting respiratory rates of <1% maximum. These results demonstrate that the sensitivity of muscle to ADP varies over a wide range in relation to contractile state and cellular energy charge, providing evidence that enzymatic coupling of energy transfer within skeletal muscle becomes more efficient in the working state. • Keywords: Bioenergetics, Blebbistatin, Creatine kinase, Myosin-ATPase, N-benzyltoluene sulfonamide, Skeletal muscle contraction
• O2k-Network Lab: US_NC Greenville_Neufer PD, CA Toronto Perry CG, US_NC_Greenville_Anderson EJ, CA_Antigonish_Kane DA
Labels: MiParea: Respiration
Organism: Human, Rat
Tissue;cell: Skeletal muscle
Preparation: Permeabilized tissue
Enzyme: Complex I, Complex V;ATP synthase
Regulation: ADP, Inhibitor, Temperature
Pathway: N HRR: Oxygraph-2k
Blebbistatin