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At least 19 recordsLinked to original sources
PubMed1988

Regional creatine kinase, adenylate kinase, and lactate dehydrogenase in normal canine brain.

Following acute stroke, creatine kinase and other enzymes are released into the cerebrospinal fluid and blood from injured brain tissue. To determine whether regional differences in brain enzyme activity might exist and therefore affect the amount of enzyme released, we quantified the levels of creatine kinase, adenylate kinase, and lactate dehydrogenase in 12 regions of normal canine brain (n = 4). Adenylate kinase activity varied the least among regions (49 +/- 7 units/g), followed by lactate dehydrogenase activity (122 +/- 28 units/g). The pattern for both adenylate kinase and lactate dehydrogenase was higher activity in predominantly gray matter areas, lower activity in white matter, and intermediate activity in mixed regions. The distribution of creatine kinase brain isoenzyme and mitochondrial creatine kinase in canine brain was less predictable, showing wider variations among regions (isoenzyme, 462 +/- 116 units/g; mitochondrial, 42 +/- 20 units/g). Even cerebral gray matter demonstrated substantial regional variations in creatine kinase brain isoenzyme, ranging from 606 units/g in the parietal cortex to 329 units/g in the temporal cortex. We conclude that the content of creatine kinase brain isoenzyme varies more than twofold among areas of brain. This regional variation may be important in the interpretation of creatine kinase brain isoenzyme measurements in cerebrospinal fluid and serum used to assess neurologic injury following stroke.

Adenylate Kinase↗
PubMed1980

Uridine kinase, adenylate kinase, and guanase in human lung tumors.

In pulmonary neoplasms, the uridine kinase concentration was higher (2- to 20-fold) than in the noninvolved lung portions of each of the 12 subjects studied. The extent of elevation of uridine kinase in the different tumors showed a significant positive correlation with the rises (1.5- to 30-fold) in thymidine kinase, suggesting that neoplastic transformation in human lung involved coordinated increases in the capacity for the reutilization of different nucleoside phosphates. Adenylate kinase was always at lower levels in neoplasms compared to noninvolved areas of the same lung, and the extent of this loss in the different tumors correlated inversely with the gain in uridine kinase and thymidine kinase. Normal fetal human lung was also deficient in adenylate kinase, while its uridine kinase and thymidine kinase (and also guanase) activities were above the adult levels. The guanase activities of the different neoplasms, unrelated to their uridine kinase or thymidine kinase content, correlated with the activities in the subjects' noninvolved lung. These individual differences were much more striking than those between the neoplastic and control samples. Variations in guanase activity thus appear to be "random," whereas observations on the three other enzymes attest to the orderly nature of biochemical differences among individual tumors and between normal and neoplastic lung.

Adenocarcinoma↗
PubMed1996

Adjustment of K' for the creatine kinase, adenylate kinase and ATP hydrolysis equilibria to varying temperature and ionic strength.

Comparative physiologists and biochemists working with tissues at varying temperatures and ionic strength are required to adjust apparent equilibrium constants (K') of biochemical reactions to the experimental conditions prior to calculating cytosolic bioenergetic parameters (transformed Gibbs free energy of formation, DeltafG' ATP; cytosolic phosphorylation ratio, [ATP]/[ADP][Pi]; [phosphocreatine]: [orthophosphate] ratio [PCr]/[Pi]) and kinetic parameters (free [ADP], [Pi] and [AMP]). The present study shows how to adjust both K' and the equilibrium constants of reference reactions (Kref) of creatine kinase (ATP: creatine N-phosphotransferase; EC 2.7.3.2), adenylate kinase (ATP:AMP phosphotransferase; EC 2.7.4.3) and adenosinetriphosphatase (ATP phosphohydrolase; EC 3.6.1.3) to temperature and ionic strength. This information, together with our previous study showing how to adjust equilibria to varying pH and pMg, is vital for the quantification of organ and tissue bioenergetics of ectotherms and endotherms under physiological conditions.

Adenosine Triphosphate↗
PubMed1995

Adjustment of K' to varying pH and pMg for the creatine kinase, adenylate kinase and ATP hydrolysis equilibria permitting quantitative bioenergetic assessment.

Physiologists and biochemists frequently ignore the importance of adjusting equilibrium constants to the ionic conditions of the cell prior to calculating a number of bioenergetic and kinetic parameters. The present study examines the effect of pH and free magnesium levels (free [Mg2+]) on the apparent equilibrium constants (K') of creatine kinase (ATP: creatine N-phosphotransferase; EC 2.7.3.2), adenylate kinase (ATP:AMP phosphotransferase; EC 2.7.4.3) and adenosinetriphosphatase (ATP phosphohydrolase; EC 3.6.1.3) reactions. We show how K' can be calculated using the equilibrium constant of a specified chemical reaction (Kref) and the appropriate acid-dissociation and Mg(2+)-binding constants at an ionic strength (I) of 0.25 mol l-1 and 38 degrees C. Substituting the experimentally determined intracellular pH and free [Mg2+] into the equation containing a known Kref and two variables, pH and free [Mg2+], enables K' to be calculated at the experimental ionic conditions. Knowledge of K' permits calculation of cytosolic phosphorylation ratio ([ATP]/[ADP][Pi]), cytosolic free [ADP], free [AMP], standard transformed Gibbs energy of formation (delta fG' degrees ATP) and the transformed Gibbs energy of the system (delta fG' ATP) for the biological system. Such information is vital for the quantification of organ and tissue bioenergetics under physiological and pathophysiological conditions.

Adenosine Triphosphate↗
PubMed1983

Simple and fast purification of Escherichia coli adenylate kinase.

Adenylate kinase from E. coli (strains CR341 and CR341 T28, a temperature-sensitive mutant) was purified by a two-step chromatographic procedure. The enzyme from crude extracts of both mutant and parent strain was bound to blue-Sepharose at pH 7.5, thereafter specifically eluted with 0.05 mM P1,P5-di(adenosine-5')pentaphosphate. A second chromatography on Sephadex G-100 yielded pure enzyme. E. coli adenylate kinase was strongly inhibited by P1,P5-di(adenosine-5')pentaphosphate (Ki 0.6 microM for adenylate kinase of strain CR341 and 2.1 microM in the case of mutant enzyme). After denaturation in 6 M guanidinium hydrochloride both mutant and parent adenylate kinase returned rapidly to the native, active state by dilution of guanidinium hydrochloride.

Adenylate Kinase↗
PubMed1991

Role of leucine 66 in the asymmetric recognition of substrates in chicken muscle adenylate kinase.

Adenylate kinase has two distinct binding sites for nucleotide substrates, MgATP and AMP. To identify the location of the site that specifically interacts with the adenine ring of AMP, we have substituted Ala, Gly, Val, Gln, and Trp for Leu66 of the recombinant chicken muscle enzyme by site-directed mutagenesis. All the purified Leu66 mutant enzymes exhibited an essentially identical circular dichroism spectrum and had thermal stabilities similar to the wild-type enzyme. Steady state kinetic analysis showed that the Leu66 mutant enzymes have significantly decreased Vmax values and markedly large Km values only for AMP. These results show that the binding site for the adenine ring of AMP in adenylate kinase is presumably located close to Leu66, which is invariant in all the enzymes so far sequenced. Significant inhibition of activities of the mutant enzymes and quenching of the Trp66 fluorescence by substrates suggest that in some Leu66 mutant enzymes, MgATP also binds to the AMP-binding site. Thus, Leu66 of adenylate kinase might play a role in the asymmetric recognition of the adenine ring of AMP from that of MgATP. Furthermore, the hydrophobicity of the residue at position 66 appears to be important for the positive cooperativity of substrate binding.

Adenosine Monophosphate↗
PubMed1982

Location and properties of two isoenzymes of cardiac adenylate kinase.

Adenylate kinase catalyses the equilibrium 2ADP = ATP + AMP. There are two isoenzymes of adenylate kinase in bovine ventricular tissue, one cytoplasmic, the other mitochondrial. Mitochondrial subfractionation locates this isoenzyme between the mitochondrial membranes with fatty acid-CoA ligase. The cytoplasmic and mitochondrial isoenzymes are distributed in ratio 3:2, and both forms were purified to homogeneity. They differ principally by charge, Km values for ATP, ADP and AMP, pH-stability and -activity profiles, and susceptibility to the inhibitor adenosine pentaphosphoadenosine. The forward and reverse reactions show similar energies of activation for the cytoplasmic enzyme, but differ for the mitochondrial enzyme. The molecular weights are indistinguishable. An integrated mechanism is formulated whereby one isoenzyme suppresses the activation of fatty acid and the other enhances carbohydrate utilization in hypoxic myocytes.

Adenine Nucleotides↗
PubMed1983

The levels of creatine kinase and adenylate kinase in the plasma of dystrophic chickens reflect the rates of loss of these enzymes from the circulation.

The rates of loss of adenylate kinase and creatine kinase from the circulation after intravenous injection of homogenous chicken skeletal muscle enzymes were examined to determine the role of plasma clearance rates in determining the plasma levels of these enzymes in normal and dystrophic chickens. The rapid clearance of adenylate kinase activity (average half-life of 5 min) and the slower biphasic clearance of creatine kinase activity (average half-lives of 0.95 and 11 hr) are consistent with the elevation of creatine kinase but not adenylate kinase in the blood plasma of dystrophic chickens compared to normal chickens. The rates of clearance of these enzymes were similar in normal chickens compared to dystrophic chickens. Radioiodinated enzymes were cleared at similar, but slightly more rapid rates than the loss of enzyme activity. The loss of adenylate kinase activity from the circulation may be due in part to inactivation since adenylate kinase activity is rapidly inactivated in serum in vitro, and because no increase in adenylate kinase activity is observed in the most specific sites of clearance of the radioiodinated enzyme, the liver and spleen. The comparison of enzyme activities in press juices to the activities in high-ionic-strength homogenates of muscle tissue from normal and dystrophic muscle, indicates that adenylate kinase activity is not associated with intracellular structures to the extent that would prohibit release from dystrophic muscle tissue. These results, and those presented previously with regard to plasma levels and clearance rates of AMP aminohydrolase and pyruvate kinase in normal and dystrophic chickens (11) support our hypothesis that the rates of loss of muscle enzyme activities from the circulation are important in determining the circulating levels of muscle enzymes in dystrophic chickens. Furthermore, from the measurement of plasma levels and clearance rates of creatine kinase, it was estimated that the efflux rate of creatine kinase from dystrophic muscle tissue is 2.0% of the total breast muscle creatine kinase per day.

Adenylate Kinase↗
PubMed1986

Adenylate kinase increases adenylate cyclase activity in membranes from rat lung.

The adenylate cyclase activity of membranes prepared from rat lung, measured under standard assay conditions, was markedly increased by the presence of a crude supernatant fraction prepared from rat lung, liver, or brain. This was not due to an increase in the initial rate of cyclic AMP (cAMP) synthesis, but to the maintenance of a constant rate of cAMP synthesis for periods of at least 10 min. After incubating lung membranes in the cyclase reaction mixture until cAMP synthesis had virtually ceased (10 min), the addition of alpha-(32P)-ATP caused a marked increase in the activity of the enzyme. This was the only component of the original reaction mixture that supported re-initiation of cAMP synthesis. Re-initiation also occurred when supernatant was added. This implies that substrate depletion occurs in the presence of membranes and that lung supernatant can catalyze rapid regeneration of substrate. Chromatographic analysis confirmed that ATP was rapidly hydrolyzed to AMP in the presence of the membranes, that this rapid destruction of ATP did not occur when supernatant was present, and that ATP was resynthesized from AMP when supernatant was added to a reaction mixture in which most of the ATP initially present had been destroyed. The effects of supernatant were mimicked by commercially available adenylate kinase. Addition of adenylate kinase did not affect adenylate cyclase activity measured in membranes prepared from brain, heart, or kidney, suggesting that lung membranes may contain more nucleotide pyrophosphatase and/or less endogenous adenylate kinase activity. Studies of soluble factors that affect adenylate cyclase must carefully control for differential substrate depletion in the presence and absence of tissue extracts.

Adenylate Kinase↗
PubMedDate not supplied

Interaction of creatine kinase and adenylate kinase systems in muscle cells.

Elsewhere in this book the important role of creatine kinase and its metabolites in high energy phosphate metabolism and transport in muscle cells has been reviewed. The emphasis of this review article is mainly on the compartmentalized catalytic activity of adenylate kinase in relation to creatine kinase isoenzymes, and other enzymes of energy production and utilization processes in muscle cells. At present the role of adenylate kinase is considered simply to equilibrate the stores of adenine nucleotides. Recent studies by us and others, however, suggest an entirely new view of the metabolic importance of adenylate kinase in muscle function. This view offers a closer interaction between adenylate kinase and creatine kinase, in the process of energy production (at mitochondrial and glycolytic sites), and energy utilization (at myofibrillar sites and perhaps other sites such as sarcoplasmic reticular, sarcolemmal membrane, etc.), thus being an integral part of the high energy phosphate transport system. This review article opens up the opportunity to further examine the metabolism of adenine nucleotides and their fluxes through the adenylate kinase system in intact muscle cells. Using an intact system, having a preserved integrity of their compartmentalized enzymes and substrates, is essential in clarifying the exact role of adenylate kinase in high energy phosphate metabolism in muscle cells.

Adenine Nucleotides↗
PubMed2006

Activation energy of catalysis-related domain motion in E. coli adenylate kinase.

Adenylate kinase from E. coli (AKeco), folded into domains CORE, AMPbd, and LID, catalyzes the reaction AMP + ATP <--> 2ADP. Previous X-ray crystallography and optical solution methods showed that the domains AMPbd and LID, and the conserved P-loop, execute large-amplitude catalysis-related motions. We used (15)N NMR spin relaxation methods to find that the simplified model-free (MF) analysis does not, whereas our general Slowly Relaxing Local Structure analysis does, detect catalytic domain motion. SRLS set for the first time the correlation time for domain motion at tau(L)perpendicular = 8.2 ns, to be compared with tau(m) = 15.1 ns for global tumbling. These results were obtained at 303 K. Herein we conduct a temperature-dependent investigation of tau(L)perpendicular and tau(m) in the range of 288-310 K. We found that the activation energy for global tumbling is Ea = 16.9 +/- 0.5 kJ/mol, the hydrodynamic volume of hydrated AKeco is 65.6 +/- 2.1 nm3, its radius is 2.50 +/- 0.03 nm, and the number of hydration layers is 1.77. The average tau(L)perpendicular value decreases from 11 ns at 288 K to 4 ns at 310 K, with activation energies of 29.7 +/- 3.3, 32.1 +/- 4.3, and 30.4 +/- 4.3 kJ/mol for the domains AMPbd and LID, and the catalytic P-loop, respectively. These values are two-to-three times smaller than typical activation energies of enzymatic reactions. Hence kinase catalysis appears not to be controlled by domain motion in the ligand-free enzyme. However, the latter process clearly facilitates important mechanical aspects such as steric recognition and capturing of the AMP and ATP substrates, their proper positioning for phosphorylation, and the release of the ADP product.

Adenylate Kinase↗
PubMed1992

Induced-fit movements in adenylate kinases.

Adenylate kinases have an M(r) around 23,000 which classifies them among the smallest phosphoryl group transferring enzymes. In order to prevent phosphoryl transfer to water, i.e. hydrolysis, these enzymes undergo induced-fit motions on substrate binding and assemble/disassemble their catalytic centres during each reaction cycle. Details of these processes have been derived from several X-ray structure analyses. The disturbance of these analyses by crystal-packing effects is discussed.

Adenylate Kinase↗
PubMed1975

A reactive arginine in adenylate kinase.

Adenylate kinase (ATP:AMP phosphotransferase, EV 2.7.4.3) from pig heart is inactivated by the specific arginyl reagent phenylglyoxal. During inactivation two molecules of phenyglyoxal are incorporated into the protein indicating the modification of one of the 11 arginine residues. The modification of other amino acids is ruled out. Chemical modification of this essential residue is prevented by high concentrations of the substrates AMP, ADP and MgATP2-. The protection of the substrates is explained by the formation of a ternary abortive enzyme-substrate complex ESS. The dissociation constants KD = [ES] - [S]/[ESS] are determined from the kinetic data of inactivation and protection.

Adenylate Kinase↗
PubMed2000

Inorganic polyphosphate kinase and adenylate kinase participate in the polyphosphate:AMP phosphotransferase activity of Escherichia coli.

Polyphosphate kinase (PPK), responsible for the processive synthesis of inorganic polyphosphate (polyP) from ATP in Escherichia coli, can transfer in reverse the terminal phosphate residue of polyP to ADP to yield ATP. PolyP also serves as a donor in a polyP:AMP phosphotransferase (PAP) activity observed in extracts of Acinetobacter johnsonii and Myxococcus xanthus. We have found that overexpression of the gene encoding PPK results in a large enhancement of PAP activity in E. coli. The PAP activity requires both PPK and adenylate kinase in equimolar amounts. PPK and adenylate kinase form a complex in the presence of polyphosphate. We discuss a phosphotransfer mechanism that involves both enzymes and enables polyP to be a phospho-donor to AMP.

Adenylate Kinase↗
PubMed2005

The crystal structure of human adenylate kinase 6: An adenylate kinase localized to the cell nucleus.

Adenylate kinases (AKs) play important roles in nucleotide metabolism in all organisms and in cellular energetics by means of phosphotransfer networks in eukaryotes. The crystal structure of a human AK named AK6 was determined by in-house sulfur single-wavelength anomalous dispersion phasing methods and refined to 2.0-A resolution with a free R factor of 21.8%. Sequence analyses revealed that human AK6 belongs to a distinct subfamily of AKs present in all eukaryotic organisms sequenced so far. Enzymatic assays show that human AK6 has properties similar with other AKs, particularly with AK5. Fluorescence microscopy showed that human AK6 is localized predominantly to the nucleus of HeLa cells. The identification of a nuclear-localized AK sheds light on nucleotide metabolism in the nucleus and the energetic communication between mitochondria and nucleus by means of phosphotransfer networks.

Active Transport, Cell Nucleus↗
PubMed2002

Activation time of myocardial oxidative phosphorylation in creatine kinase and adenylate kinase knockout mice.

Our goal was to determine whether mice genetically altered to lack either creatine kinase (M/MtCK(-/-)) or adenylate kinase (AK(-/-)) show altered properties in the dynamic regulation of myocardial oxygen consumption (MVO(2)). We measured contractile function, oxygen consumption, and the mean response time of oxygen consumption to a step increase in heart rate [i.e., mitochondrial response time (t(mito))] in isolated Langendorff-perfused hearts from wild-type (n = 6), M/MtCK(-/-) (n = 6), and AK(-/-) (n = 4) mice. Left ventricular developed pressure was higher in M/MtCK(-/-) hearts (88.2 +/- 6.8 mmHg) and lower in AK(-/-) hearts (46.7 +/- 9.4 mmHg) compared with wild-type hearts (60.7 +/- 10.1 mmHg) at the basal pacing rate. Developed pressure fell slightly when heart rate was increased in all three groups. Basal MVO(2) at 300 beats/min was 19.1 +/- 2.4, 19.4 +/- 1.5, and 16.3 +/- 1.9 micromol x min(-1) x g dry wt(-1) for M/MtCK(-/-), AK(-/-), and wild type, respectively, which increased to 25.5 +/- 3.7, 25.4 +/- 2.6, and 22.0 +/- 2.6 micromol. min(-1) x g(-1), when heart rate was increased to 400 beats/min. The t(mito) was significantly faster in M/MtCK(-/-) hearts: 3.0 +/- 0.3 versus 7.3 +/- 0.6 and 8.0 +/- 0.4 s for M/MtCK(-/-), AK(-/-), and wild-type hearts, respectively. Our results demonstrate that MVO(2) of M/MtCK(-/-) hearts adapts more quickly to an increase in heart rate and thereby support the hypothesis that creatine kinase acts as an energy buffer in the cytosol, which delays the energy-related signal between sites of ATP hydrolysis and mitochondria.

Adenosine Triphosphate↗