title: High K(m) of oxidative phosphorylation for ADP in skinned muscle fibers: where does it stem from?
abstract: Mitochondria in saponin-skinned cardiac fiber bundles were reported to have an order of magnitude lower apparent affinity to ADP than isolated mitochondria. Although ADP was measured outside the bundles, it was thought that the low affinity was not caused by diffusion gradients because of relatively short diffusion distances. Here we test the hypothesis that considerable ADP diffusion gradients exist and can be diminished by increasing the intrafiber ADP production rate. We increased the ADP-producing activity in rat heart skinned fiber bundles by incubating with 100 IU/ml yeast hexokinase and glucose. Consequently, we observed a significant decrease of the apparent Michaelis constant (K(m)) to ADP of the respiration rate of bundles from 216 +/- 59 to 50 +/- 9 microM. Fitting the results with a mathematical model, we estimated the K(m) of mitochondria in the bundles to be 25 microM. We conclude that the affinity to ADP of in situ mitochondria in heart is of the same order of magnitude as that of isolated mitochondria.
authors.affiliation: Laboratory for Physiology, Institute for Cardiovascular Research, Vrije Universiteit Medical Center 1081 BT Amsterdam, The Netherlands. kongas@ioc.ee
authors.forename: Olav
authors.initials: O
authors.lastname: Kongas
authors.forename: Tai L
authors.initials: TL
authors.lastname: Yuen
authors.forename: Marijke J
authors.initials: MJ
authors.lastname: Wagner
authors.forename: Johannes H G M
authors.initials: JH
authors.lastname: Van Beek
authors.forename: Klaas
authors.initials: K
authors.lastname: Krab
mesh_terms: D000244:Adenosine Diphosphate / Q000378:metabolism*; D000818:Animals; D002451:Cell Compartmentation; D004058:Diffusion; D005260:Female; D005947:Glucose / Q000378:metabolism; D006593:Hexokinase / Q000378:metabolism; D066298:In Vitro Techniques; D008297:Male; D008929:Mitochondria, Heart / Q000378:metabolism; D008954:Models, Biological; D018485:Muscle Fibers, Skeletal / Q000378:metabolism*; D018482:Muscle, Skeletal / Q000378:metabolism; D009206:Myocardium / Q000378:metabolism; D010085:Oxidative Phosphorylation; D010101:Oxygen Consumption; D051381:Rats; D017208:Rats, Wistar; D012441:Saccharomyces cerevisiae
chemical_list: D000244:Adenosine Diphosphate; D006593:Hexokinase; D005947:Glucose