scholarly journals Reactions of 14C-ADP and 14C-ATP With Washed Platelets From Rabbits

Blood ◽  
1971 ◽  
Vol 37 (5) ◽  
pp. 542-555 ◽  
Author(s):  
M. A. GUCCIONE ◽  
M. A. PACKHAM ◽  
R. L. KINLOUGH-RATHBONE ◽  
J. F. MUSTARD
Keyword(s):  
Atpase Activity ◽  
Adenylate Kinase ◽  
Kinase Activity ◽  
Major Change ◽  
High Energy ◽  
High Energy Phosphate ◽  
Low Concentrations ◽  
Rabbit Platelets ◽  
Calcium And Magnesium ◽  
Induced Aggregation

Abstract Using suspensions of washed rabbit platelets that react to low concentrations of ADP in the same way as platelets in plasma, it has been found that the major change in added 14C-ADP is its conversion to 14C-ATP, catalyzed by nucleoside diphosphokinase at the platelet membrane. This enzyme is readily released from the platelets but the rate and extent of 14C-ADP to 14C-ATP conversion are greater in the presence of platelets than in the suspending fluid from which they have been removed. The platelets must provide the source of high-energy phosphate for the reaction, either from ATP released into the suspending fluid, or by the transfer of high-energy phosphate across the membrane. Thrombin stimulates the conversion, probably because it releases ATP from the platelets. Calcium and magnesium stimulate the conversion, whereas two of the inhibitors of ADP-induced aggregation (AMP and parachloromercuribenzenesulfonate) inhibit it. The ATPase activity of these platelets is low and it is not released into the suspending fluid. No adenylate kinase activity was demonstrable. The loss of high energy phosphate from the platelets that occurs during the conversion of 14C-ADP to 14C-ATP may be involved in the initiation of their response to ADP.

Cardiac force and high-energy phosphates under metabolic inhibition in four ectothermic vertebrates

1996 ◽  
Vol 271 (4) ◽  
pp. R946-R954 ◽  
Author(s):  
T. Hartmund ◽  
H. Gesser
Keyword(s):  
Creatine Kinase ◽  
Atpase Activity ◽  
Kinase Activity ◽  
Energy State ◽  
High Energy ◽  
Creatine Kinase Activity ◽  
The Other ◽  
Freshwater Turtle ◽  
High Energy Phosphates ◽  
Isometric Twitch

Isometric twitch tension of ventricular preparations stimulated at 0.2 Hz fell over 30 min of anoxia by a fraction decreasing in the order rainbow trout, cod, eel, and freshwater turtle. Drops in the estimated cytoplasmic energy state were related to larger tension losses for trout than for the other species, possibly due to larger changes in free phosphate. Anoxic energy degradation was slower for turtle than for the other species. Anoxia combined with glycolytic inhibition (1 mmol/l iodoacetate) enhanced the decrease in twitch tension for a drop in energy state and enlarged the increase in ADP/ATP relative to that in creatine/phosphocreatine to an extent inversely related to the creatine kinase activity. Furthermore, it increased resting tension to an extent possibly related to myosin-adenosinetriphosphatase (ATPase) activity and lowered the content of phosphorylated adenylates in trout and turtle myocardium. The results indicate that species differences in performance of the metabolically challenged myocardium depend on energy-degrading processes, e.g., myosin-ATPase activity, phosphate release, creatine kinase activity, and efflux/degradation of ADP and AMP, and that glycolysis offers protection due to its cytoplasmic localization.

scholarly journals LptB2FG is an ABC transporter with Adenylate Kinase activity regulated by LptC/A recruitment

2021 ◽  
Author(s):  
Tiago Baeta ◽  
Karine Giandoreggio-Barranco ◽  
Isabel Ayala ◽  
Elisabete CCM Moura ◽  
Paola Sperandeo ◽  
...  
Keyword(s):  
Atpase Activity ◽  
Abc Transporter ◽  
Adenylate Kinase ◽  
Kinase Activity ◽  
Cell Envelope ◽  
Transmembrane Helix ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Entire Cell ◽  
Lipid Environment

Lipopolysaccharide (LPS) is an essential glycolipid covering the surface of gram-negative bacteria. Its transport involves a dedicated 7 protein transporter system, the Lpt machinery, that physically spans the entire cell envelope. LptB2FG complex is an ABC transporter that hydrolyses Adenosine Triphosphate (ATP) to extract LPS from the inner membrane (IM). LptB2FG was extracted directly from IM with its original lipid environment by Styrene-Maleic acids polymers(SMA). SMA-LptB2FG in nanodiscs displays ATPase activity and a previously uncharacterized Adenylate Kinase (AK) activity. It catalyzes phosphotransfer between two ADP molecules to generate ATP and AMP. ATPase and AK activities of LptB2FG are both stimulated by the interaction on the periplasmic side with LptC and LptA partners and inhibited by the presence of LptC transmembrane helix. Isolated ATPase module (LptB) has weak AK activity in absence of LptF and LptG, and one mutation, that weakens affinity for ADP, has AK activity similar to that of fully assembled complex. LptB2FG is thus capable of producing ATP from ADP depending on the assembly of the Lpt bridge and the AK activity might be important to ensure efficient LPS transport in fully assembled Lpt system.

Factors Influencing the Deaggregation of Human and Rabbit Platelets

1983 ◽  
Vol 49 (03) ◽  
pp. 162-167 ◽  
Author(s):  
R L Kinlough-Rathbone ◽  
J F Mustard ◽  
D W Perry ◽  
E Dejana ◽  
J-P Cazenave ◽  
...  
Keyword(s):  
Human Platelets ◽  
Release Reaction ◽  
Factors Influencing ◽  
Platelet Adherence ◽  
Activated Platelets ◽  
Fibrinogen Binding ◽  
Low Concentrations ◽  
Rabbit Platelets ◽  
High Concentrations ◽  
Induced Aggregation

SummaryThe mechanisms involved in platelet deaggregation are unclear. Washed platelets from rabbits or humans aggregated by ADP can be deaggregated by EDTA or PGI2 if the release reaction has not occurred; during deaggregation 125I-fibrinogen dissociates from the platelets. Human platelets suspended in a medium without calcium undergo the release reaction during ADP-induced aggregation; EDTA, PGE, or PGI2 do not deaggregate these platelets although EDTA displaces much of the 125I-fibrinogen that associates with them during aggregation. Rabbit platelets aggregated by low concentrations of releaseinducing stimuli (sodium arachidonate, collagen or thrombin) can be deaggregated by EDTA, PGI2 or PGE1 and 125I-fibrinogen dissociates from them; with high concentrations of collagen or thrombin, deaggregation and dissociation of l25I-fibrinogen is slower. Human platelets that have undergone the release reaction in response to thrombin, collagen or a combination of sodium arachidonate and ADP are not readily deaggregated by EDTA or PGE1. Since aggregation and fibrinogen binding involving the glycoprotein IIb/IIIa complex are readily reversed by EDTA, and since Ca2+ is required for thrombospondin binding to activated platelets, there may be a third type of platelet-platelet adherence that is not disrupted by EDTA; this type of binding plays a greater role with human than with rabbit platelets.

Effects of Tris on Responses of Human and Rabbit Platelets to Aggregating Agents

1984 ◽  
Vol 51 (02) ◽  
pp. 140-144 ◽  
Author(s):  
M A Packham ◽  
M A Guccione ◽  
M Nina ◽  
R L Kinlough-Rathbone ◽  
J F Mustard
Keyword(s):  
Primary Amines ◽  
Amino Sugars ◽  
Ionophore A23187 ◽  
Tyrode Solution ◽  
Low Concentrations ◽  
Rabbit Platelets ◽  
Induced Aggregation ◽  
Inhibition Of Thrombin

SummaryDespite reports that Tris [tris(hydroxymethyl)aminomethane] affects platelets, it is often used to buffer suspending media. Human or rabbit platelets were washed and resuspended in Tyrode solution containing apyrase and 0.35% albumin. Addition of 15 mM Tris partially inhibited primary aggregation induced by 10 μM ADP and inhibited aggregation and release of 14C- serotonin from prelabelled platelets stimulated with low concentrations of thrombin (0.05-0.2 U/mL), or collagen. Platelets resuspended in 15 mM Tris, 0.15 M NaCl, 0.35% albumin, pH 7.5, did not aggregate in response to 10 μM ADP whereas platelets in Tyrode-albumin aggregated extensively. Ca2+ (5 mM) did not overcome the inhibition of thrombin-induced aggregation. Tris (15 or 1.5 mM) potentiated aggregation and release induced by sodium arachidonate (20-50 μM) or the ionophore A23187 (0.6-1 μM). Pretreatment of platelets with aspirin did not prevent potentiation by A23187, indicating that it is not mediated through activation of the arachidonate pathway. The inhibitory and potentiating effects of Tris are similar to those of amino sugars, lysine, arginine and primary amines such as methylamine and cadaverine, and may represent general effects of amines on platelets. Potentiation of the effects of some aggregating agents and inhibition of others re-emphasizes the concept that there are several different mechanisms through which aggregation can occur. Tris-based buffers are unsuitable for platelet suspending media and their use as solvents for aggregating agents or inhibitors should be limited.

The intact CFTR protein mediates ATPase rather than adenylate kinase activity

2008 ◽  
Vol 412 (2) ◽  
pp. 315-321 ◽  
Author(s):  
Mohabir Ramjeesingh ◽  
Francisca Ugwu ◽  
Fiona L. L. Stratford ◽  
Ling-Jun Huan ◽  
Canhui Li ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Atpase Activity ◽  
Mechanism Of Action ◽  
Adenylate Kinase ◽  
Kinase Activity ◽  
Residual Activity ◽  
Atp Binding ◽  
Cftr Protein ◽  
The Individual ◽  
Atp Binding And Hydrolysis

The two NBDs (nucleotide-binding domains) of ABC (ATP-binding-cassette) proteins function in a complex to mediate ATPase activity and this activity has been linked to their regulated transport activity. A similar model has been proposed for CFTR (cystic fibrosis transmembrane conductance regulator), the chloride channel defective in cystic fibrosis, wherein ATP binding and hydrolysis regulate the channel gate. Recently, it was shown that the individual NBDs isolated from CFTR primarily mediate adenylate kinase activity, raising the possibility that this activity may also contribute to gating of the CFTR channel. However, this present study shows that whereas the isolated NBDs exhibit adenylate kinase activity, the full-length purified and reconstituted CFTR protein functions as an ATPase, arguing that the enzymatic activity of the NBDs is dependent on their molecular context and appropriate domain–domain assembly. As expected, the disease-causing mutant bearing a mutation in the ABC signature motif, CFTR-G551D, exhibited a markedly reduced ATPase activity. Furthermore, mutation of the putative catalytic base in CFTR caused a reduction in ATPase activity, with the CFTR-E1371Q mutant supporting a low level of residual activity. Neither of these mutants exhibited detectable adenylate kinase activity. Together, these findings support the concept that the molecular mechanism of action of CFTR is dependent on ATP binding and hydrolysis, and that the structure of prokaryotic ABC ATPases provide a useful template for understanding their mechanism of action.

Adenylate kinase inhibition by adenosine 5'-monophosphate and fluoride in the determination of creatine kinase activity.

1978 ◽  
Vol 24 (3) ◽  
pp. 498-501 ◽  
Author(s):  
F Meiattini ◽  
G Giannini ◽  
P Tarli
Keyword(s):  
Creatine Kinase ◽  
Adenylate Kinase ◽  
Kinase Activity ◽  
Clinical Chemistry ◽  
German Society ◽  
Significant Loss ◽  
Creatine Kinase Activity ◽  
Low Concentrations ◽  
Diadenosine Pentaphosphate

Abstract The current methods for the determination of creatine kinase (EC 2.7.3.2) activity are derived from Oliver's method, in which AMP is used to decrease interference by adenylate kinase (EC 2.7.4.3). Recently, Szasz et al. and Rosano et al. described methods in which diadenosine pentaphosphate and fluoride, respectively, are used to reduce this interference. However, diadenosine pentaphosphate does not sufficiently inhibit such activity of hepatic origin, while fluoride alone can only inhibit it at concentrations at which the fluoride tends to precipitate as MgF2. Finally, Szasz et al., the Committee on Enzymes of the Scandinavian Society for Clinical Chemistry and Clinical Physiology, and the German Society for Clinical Chemistry have proposed methods in which both AMP and diadenosine pentaphosphate are used to inhibit adenylate kinase. We have found that by using low concentrations of AMP and fluoride together, we can greatly diminish this interference without significant loss of creatine kinase activity and with no precipitation of MgF2.

scholarly journals Characterization of the Mg2+-activated ATPase activity in smooth-muscle membranes. NADH oxidase and adenylate kinase interfere with the NADH-coupled enzyme assay

1988 ◽  
Vol 250 (2) ◽  
pp. 579-588 ◽  
Author(s):  
L Missiaen ◽  
F Wuytack ◽  
R Casteels
Keyword(s):  
Smooth Muscle ◽  
Atpase Activity ◽  
Adenylate Kinase ◽  
Oxidase Activity ◽  
Kinase Activity ◽  
Nadh Oxidase ◽  
Taenia Coli ◽  
Ear Artery ◽  
Rabbit Ear ◽  
Nadh Oxidase Activity

The apparent Mg2+-activated ATPase activity measured by the continuous NADH-coupled enzyme assay was studied in a number of microsomal preparations obtained from smooth muscle of the myometrium from pregnant or 17 beta-oestradiol-pretreated rats, the bovine aorta, the guinea-pig taenia coli, the rabbit ear artery and pig antrum. It was shown that this ATPase assay is prone to the effects of a number of artefacts that are tissue-dependent. The apparent Mg2+-ATPase activity in microsomes (microsomal fractions) from myometrium, aorta and taenia coli declines non-linearly during the assay. Its initial high rate gradually diminishes over 15-60 min, depending on the type of smooth muscle, to a constant value. This decline depends on the presence of ATP and can be partially prevented by concanavalin A. The non-linearity is limited in microsomes from rabbit ear artery. In microsomes from antrum the apparent Mg2+-ATPase activity actually increases with time, albeit gradually. Storage on ice of the microsomes of the aorta, and especially of myometrium of pregnant rats and of taenia coli, is accompanied over a few hours after their preparation by a gradual suppression of the component of the Mg2+-ATPase activity that is inhibited by ATP. The Mg2+-ATPase activity in microsomes from antrum remains constant. NADH oxidase activity accounts for 10% of the Mg2+-ATPase activity in microsomes from stomach smooth muscle. The apparent initial non-linearity of the Mg2+-ATPase activity in that tissue is due to a time-dependent decrease of a rotenone-sensitive NADH oxidase activity. The adenylate kinase activity, as deduced from the effect of the adenylate kinase inhibitor P1,P5-di(adenosine-5′) pentaphosphate, could account for 45.0, 35.0 and 31.0% respectively of the Mg2+-ATPase activity in microsomes from stomach, myometrium and aorta. No adenylate kinase activity could be detected in microsomes from ear artery and taenia coli. When microsomes from stomach smooth muscle were separated on a sucrose gradient, the contribution of adenylate kinase and NADH oxidase to the Mg2+-ATPase activity was most pronounced in the higher-density fractions. Part of the NADH oxidase activity and of the Mg2+-ATPase activity, and most of the adenylate kinase activity, are not sedimented at 224000 gmax. for 30 min and may therefore be present as soluble enzymes.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals ATP Metabolism in Pyruvate Kinase Deficient Erythrocytes

Blood ◽  
1967 ◽  
Vol 30 (5) ◽  
pp. 576-586 ◽  
Author(s):  
JEREMIAH J. TWOMEY ◽  
FLOYD B. O’NEAL ◽  
CLARENCE P. ALFREY ◽  
ROBERT H. MOSER ◽  
Wilma J. Hudson
Keyword(s):  
Atpase Activity ◽  
Pyruvate Kinase ◽  
Aerobic Glycolysis ◽  
High Energy ◽  
Red Cells ◽  
Red Cell ◽  
High Energy Phosphate ◽  
Atp Metabolism ◽  
Atp Generation

Abstract Elevated levels of ATP were observed in pyruvate kinase deficient red cells from 5 patients despite increased ATP requirements because of increased ATPase activity. Increased red cell ATPase activity was associated with an excess of intracellular sodium. Residual aerobic glycolysis in reticulocytes did not contribute significantly toward ATP generation in these patients’ red cells. Their hemolytic anemia cannot be ascribed to a lack of high energy phosphate. Conversely, hemolysis in vitro appears to require a high energy phosphate dependent metabolic process.

Strategies and protocols for clinical 31 P research in the heart and brain

1990 ◽  
Vol 333 (1632) ◽  
pp. 531-544 ◽  
Keyword(s):  
Nmr Relaxation ◽  
Patient Positioning ◽  
High Energy ◽  
Spectral Distortion ◽  
High Energy Phosphate ◽  
Low Concentrations ◽  
Field Uniformity ◽  
Critical Limits ◽  
Volume Size

Although in vivo 31 P NMR permits unique measurements of the body’s stores of high energy phosphate metabolites, their low concentrations place critical limits and demands on spatial resolution, localization techniques, and reproducibility. We argue that optimum reliability and reproducibility of in vivo measurements of metabolite ratios and concentrations is best achieved by preselecting the minimal requirements for spatial localization suited to, and varying with, the particular organ and disease type. Strategies for measuring metabolite ratios and concentrations in the human heart and brain are discussed and demonstrated in the context of the expeditious choice of operating parameters, including localized volume size, localization technique, NMR coils, radiofrequency-field uniformity, patient positioning, and corrections for spectral distortion caused by NMR relaxation, to optimize the return of useful metabolic information in practical clinical research exams of about an hour duration.

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