Myokinase and contractile function of glycerinated muscle fibers

Matching blood flow to myocardial energy demand is vital for heart performance and recovery following ischemia. The molecular mechanisms responsible for transduction of myocardial energetic signals into reactive vasodilatation are, however, elusive. Adenylate kinase, associated with AMP signaling, is a sensitive reporter of the cellular energy state, yet the contribution of this phosphotransfer system in coupling myocardial metabolism with coronary flow has not been explored. Here, knock out of the major adenylate kinase isoform, AK1, disrupted the synchrony between inorganic phosphate P i turnover at ATP-consuming sites and ␥-ATP exchange at ATP synthesis sites, as revealed by 18 O-assisted 31 P NMR. This reduced energetic signal communication in the post-ischemic heart. AK1 gene deletion blunted vascular adenylate kinase phosphotransfer, compromised the contractility-coronary flow relationship, and precipitated inadequate coronary reflow following ischemiareperfusion. Deficit in adenylate kinase activity abrogated AMP signal generation and reduced the vascular adenylate kinase/ creatine kinase activity ratio essential for the response of metabolic sensors. The sarcolemma-associated splice variant AK1␤ facilitated adenosine production, a function lost in the absence of adenylate kinase activity. Adenosine treatment bypassed AK1 deficiency and restored post-ischemic flow to wild-type levels, achieving phenotype rescue. AK1 phosphotransfer thus transduces stress signals into adequate vascular response, providing linkage between cell bioenergetics and coronary flow.Coronary blood flow is tightly linked to myocardial oxygen consumption securing adequate oxygenation and nutrient delivery and preventing underperfusion (1-4). Mismatch between myocardial blood supply and energy metabolism precipitates heart dysfunction, leading to aberrant force-frequency relationships in acute coronary and chronic metabolic syndromes (4 -7). The no-reflow phenomenon, observed after vascular reperfusion, precipitates poor outcome (8). Despite advances made in elucidating components that regulate vascular tone, the intimate mechanisms integrating metabolic signals and securing adequate vasodilatation under stress remain partially understood (4, 9 -13).Regulation of vascular tone involves several effectors, including ATP-sensitive potassium (K ATP ) channels, adenosine and purinergic signaling, AMP-activated protein kinase (AMPK), 3 nitric oxide (NO), and neurohormonal factors (1-4, 9 -13). Intra-and extracellular nucleotide-mediated signaling pathways catalyzed by phosphotransfer enzymes may provide linkage between these otherwise distinct systems (14 -16). In particular, adenylate kinase phosphotransfer couples with K ATP channels, facilitating decoding of metabolic signals critical in adjusting excitability-dependent functions in response to demand (17, 19 -21). Adenylate kinase, with a unique property to catalyze the reaction 2ADP 7 ATPϩAMP, is a sensitive reporter of the cellular energy state, translating small changes in the ATP/AD...

Section: Discussionmentioning

confidence: 99%

Defective Metabolic Signaling in Adenylate Kinase AK1 Gene Knock-out Hearts Compromises Post-ischemic Coronary Reflow

Dzeja

Bast

Pucar

et al. 2007

“…In this regard, previous functional studies on fast-twitch muscles of mice lacking M-CK have demonstrated lack of burst activity (24) and compromised ability to sustain contraction force (43), in line with loss of preferential access to ATP via the CK reaction. Similarly, AK1 activity facilitates muscle relaxation, indicating that removal of contraction-produced ADP by AK1 and its phosphorylation to ATP takes place much faster than diffusional exchange of ADP to ATP (47). As ultra-fast local and global ATP regeneration is required to sustain contractile activity in fast-twitch muscle (24,52), this would underscore the particular importance of CK-and AK-catalyzed phosphotransfer in these muscle types.…”

Section: Figmentioning

confidence: 99%

“…AK1 also localizes on the myofibrillar I-band where it may work as an ATP regenerator to support muscle contraction (13,21,47). To determine whether this cellular AK1 and M-CK partitioning has functional significance, we compared the contractile properties of actomyosin complexes isolated from wild-type and MAK ϭ/ϭ muscles.…”

Section: Cellular Energetics In Resting Andmentioning

confidence: 99%

Impaired Intracellular Energetic Communication in Muscles from Creatine Kinase and Adenylate Kinase (M-CK/AK1) Double Knock-out Mice

Janssen

Terzic

Wieringa

et al. 2003

Previously we demonstrated that efficient coupling between cellular sites of ATP production and ATP utilization, required for optimal muscle performance, is mainly mediated by the combined activities of creatine kinase ( In tissues with high and sudden energy demand, creatine kinase (CK) 1 -and adenylate kinase (AK)-catalyzed reactions form the principal pathways securing efficient communication between the subcellular compartments responsible for production and utilization of metabolic energy (1-6).Adenylate kinases (AK, EC 2.7.4.3), an evolutionary conserved family of enzymes that catalyzes the reaction ATP ϩ AMP 7 2 ADP (7), have been implicated in cellular adenine nucleotide homeostasis (8). cDNAs for five isoforms of AK (AK1-AK5) along with the variant of AK1 (AK1␤, a membranebound form with a presumed role in cell cycle regulation) have been cloned from metabolically active tissues (9 -12). Mammalian skeletal muscle is particularly rich in AK1, the major isoform of the family (9), present in the sarcoplasm, and clustered along the myofibrillar I-band or bound as AK1␤ to membranes (13-15). By donating the energy of the ␤-phosphoryl group of ATP/ADP to the cellular energetic pool, AK isoenzymes protect cells against energy deprivation in periods of high metabolic demand (6, 16 -20). The different intracellular localizations and distinct kinetic properties of AK isoforms permit the formation of a coordinated enzymatic network for nucleotide-mediated metabolic signaling, coupling myofibrillar, nuclear, or sarcolemmal energy-dependent processes with mitochondrial energetics (21-23).Creatine kinases (CK, EC 2.7.3.2) catalyzing the reaction MgADP Ϫ ϩ CrP 2Ϫ ϩ H ϩ 7 Cr ϩ MgATP 2Ϫ belong to a smaller and evolutionary younger family of enzymes with a role in high energy phosphoryl transfer and cellular energy buffering (1,5,24). Creatine kinases are foremost found in cells with high peak demands in metabolic energy such as the brain, heart, or skeletal muscle (1, 5). In skeletal muscle, the principal CK isoform is the cytosolic isoform, MM-CK, a homodimer mainly present as a soluble protein in the cytosol and bound to the myofibrillar M-and I-bands (13) as well as to the sarcoplasmic reticulum membranes (25). Skeletal muscle also contains an additional mitochondrial CK isoform (ScCKmit), which amounts to 1-10% of the total CK activity depending on the type of muscle fiber (26,27). This CK member associates and functionally interacts with the adenine nucleotide translocator and voltage-dependent anion channel in the mitochondrial inner and outer membrane (28 -30), providing an efficient ATP export and metabolic signal reception pathway (31).AK and CK in concert with nucleoside diphosphokinase (NDPK) and the enzymes that function in the glycolytic phosphotransfer pathway form the cellular energetic infrastructure responsible for effective handling and distribution of high energy phosphoryl (ϳP) groups throughout the structured muscle environment (5,6,24,(32)(33)(34). In this network, AK-and CKmediated reactions play a...

“…Additionally, this mechanism would also help to resolve the problem created by the abundant and very catalytically active myofibrillar and cytosolic CK and AK enzyme protein molecules with the potential to bind and catalytically transform ATPase-generated ADP before it could freely diffuse from its site of generation to an ATP regenerating site (22)(23)(24)(25). Another feature of this proposed conduction system underscoring its potential thermodynamic efficiency is its capability to operate with minimal or no concentration gradient of reactants (31)(32)(33).…”

Section: Figmentioning

confidence: 99%

Suppression of Creatine Kinase-catalyzed Phosphotransfer Results in Increased Phosphoryl Transfer by Adenylate Kinase in Intact Skeletal Muscle

Dzeja

Zeleznikar

Goldberg

1996

103

The kinetics of creatine kinase (CK) and adenylate kinase (AK) activities were monitored in intact diaphragm muscle by 18 O phosphoryl oxygen exchange to assess whether these two phosphotransferases provide an interrelated function integral to high energy phosphoryl metabolism. This possibility was examined by quantitating the net rates of CK-and AK-catalyzed phosphoryl transfer in comparison to the total cellular ATP metabolic rate when CK activity in the intact diaphragm muscle was progressively inhibited by 2,4-dinitrofluorobenzene. In noncontracting muscle from untreated rats, net rates of CK-and AK-catalyzed phosphotransfer were equivalent to 88 and 7%, respectively, of the total ATP metabolic rate. These results were compared with reported 31 P NMR analyses of total creatine phosphate flux to estimate that each creatine phosphate molecule produced undergoes about 50 unidirectional CK-catalyzed phosphotransfers in transit to an ATP consumption site in the intact muscles. Graded inhibition by 2,4-dinitrofluorobenzene of intracellular CK activity by up to 98% resulted in a progressive shift in phosphotransferase catalysis from the CK to the AK system; the sum of the net rates of phosphoryl transfer by combining the increasing AK and decreasing CK activities continued to approximate the total cellular ATP metabolic rate. These results indicate that in diaphragm muscle CK and AK operate as interrelated cellular high energy phosphoryl transfer systems through which the majority of newly generated ATP is processed prior to its utilization.The view put forth by Bessman and co-workers (1, 2) that the metabolic function of creatine kinase (CK) 1 is to transfer energy-rich phosphoryls from intracellular generation to utilization sites is supported by kinetic studies in vitro and in vivo by the distinctive localization of isozymic forms of CK and their functional coupling to ATP-consuming and ATP-generating processes (for reviews see . In opposition to this view is the relatively intact physiological performance of muscle when CK activity is impaired following protracted ingestion of creatine analogs (6, 7) or deletion of the gene encoding the major cytosolic isoform of muscle CK (i.e., M-CK) (8). It is also contradicted by reports that the total unidirectional rate of CKcatalyzed transfer of phosphoryls assessed by 31 P NMR technology in intact skeletal muscle is workload-independent or even reduced with the onset of muscle contraction (9, 10).An importance in intracellular energy transfer similar to that of CK has been suggested for adenylate kinase (AK)-catalyzed phosphotransfer (4,11,12). From studies of AK kinetic behavior in intact diaphragm muscle, increases in net AK-catalyzed phosphotransfer were found to be directly proportional to the frequency of stimulated muscle contraction (12), and the stoichiometry between net AK-catalyzed phosphoryl transfer and anaerobic glycolytic ATP generation was nearly equivalent over a greater than 20-fold range of stimulated fluxes (13). The interpretation of these results wa...