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Query: UMLS:C0027960 (mole)
 21,279 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The kinetic properties of pH jump-induced phosphorylation in thylakoidal membranes of spinach chloroplasts were investigated, and the following results were obtained. 1. The pH jump-induced P incorporation proceeded linearly with time for at least 2 sec after the start of the reaction. Phosphate was incorporated mainly into ATP. The amounts of incorporation into ADP during 0.1 and 2.0 sec were 0.02 and 0.1 mole/400 moles chl, respectively. The amounts of P incorporation into ADP and the beta-position of ATP during a 2 sec reaction were less than 5 and 0.1% of the total amount of P incorporation, respectively. Even in the absence of added ADP, ATP was formed by the pH jump, but the amount was very small, i.e., less than 1% of that in the presence of a saturating amount of ADP. Formation of ATP was not enhanced by the addition of 0.1 mM AMP, instead of ADP. 2. The dependence of the rate of ATP formation, v, induced by a pH jump from 3.85 to 8.11 on the concentrations of ADP and Pi was given by v=Vopt/[1+psi1/[ADP]) (1 + psi2/[Pi)], where the values of the constants, Vopt, psi1, and psi2 were 14--20 moles/10-6 g chl/sec, 12.5-15 muM and 11-20 mM, respectively, at 0 degrees. 3. The dependence of v on the concentration of protons was given by v=Va/[1 + psi H-a/[H+,-a])-2], and v =Vb/[1 + ([H+,-b]/psiH-b)-2], in the acidic and basic phases, respectively. The values of the constants psi H-a and psi H-b were 10-5.7 and 10-7.9 M, respectively. 4. ATP formation was initiated by adding one of the substrates, ADP or Pi, at various times after after the pH jump in the presence of the other substrate. The rate decreased logarithmically with increase in the time between the pH jump and the start of the reaction. When phosphorylation was initiated by adding Pi after the pH jump in the presence of ADP, the decay constant of v was about 0.08 sec-1, which was one-third of that observed when the order of addition of ADP and Pi was reversed.
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PMID:pH jump-induced phosphorylation of adenosine diphosphate in thylakoidal membranes. 23 94

1. Membrane potential changes produced by adenosine and adenine nucleotides, acetylcholine, and vagus nerve stimulation were studied by intracellular recording in the sinus venosus of the frog, Rana pipiens. 2. Acetylcholine (ACh) released from the vagus nerve terminals evoked a slow hyperpolarization lasting several seconds in the cells of the sinus. Ionophoretic application of ACh from a micropipette produced a response which is similar in time course and amplitude to that evoked by vagus nerve stimulation. Bath application of ACh caused a steady hyperpolarization in quiescent preparations, or cessation of action potential generation in spontaneously active preparations. 3. Adenosine and adenine nucleotides produced hyperpolarizations when applied by addition to the bath or by ionophoresis from micropipettes. The hyperpolarization produced by ionophoresis of adenine compounds was somewhat slower than that produced by ACh. 4. Adenosine and the adenine nucleotides, 5'-AMP, 3'-AMP, 2'-AMP, and 5'-atp were virtually equipotent in their action. Adenosine was at least 1000-fold more potent than other purine and pyrimidine nucleosides or adenine. Both the ribose and adenine groups were important for agonist activity. 5. The concentrations of agonist required to produce half-maximal responses were estimated from dose--response curves as 3 x 10(-7) M for ACh and 2 x 10(-6) M for ATP. ACh is about 7 times more potent than ATP in producing a hyperpolarization. 6. Adenine compounds act directly upon the cardiac muscle fibres: bath or ionophoretically applied adenine compounds act even when transmitter release from nerve terminals is blocked with high (Mn2+) or when ACh receptors are blocked with atropine. 7. Adenine compounds act on the surface of the muscle fibre membrane. Analogues of adenosine which do not enter the cell are potent agonists of the receptor. An adenyl oligonucleotide too large to enter the cell was 2.6 times more potent per mole than adenosine in producing a hyperpolarization. Drugs such as dipyridamole and 6-(2-hydroxy 5-nitrobenzyl) thioguanosine, which are potent blockers of adenosine transport, potentiate the response of the sinus cells to adenosine. 8. Aminophylline and theophylline are competitive antagonists of adenosine action. The apparent Ki for aminophylline inhibition was 5 microM. 9. The response produced by adenine compounds is partly caused by an increase in the permeability of the membrane to K+. The maximum response to both ACh and adenine nucleotides approached the estimated level of EK or ECl. Replacing extracellular chloride with impermeant isethionate had no effect on responses to ACh or adenine nucleotides. The hyperpolarization was not produced by an activation of an ouabain-sensitive pump since 20 microM-ouabain had little effect on the response to adenosine. 10. The response to vagus nerve stimulation is completely blocked by 50 nM-atropine...
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PMID:Adenosine receptors in frog sinus venosus: slow inhibitory potentials produced by adenine compounds and acetylcholine. 31 61

The binding of nine aminoalkyl adenylates to isoleucyl-tRNA synthetase from Escherichia coli MRE 600 was measured and compared with the binding of the cognate amino acids. It was found that they bind rather tightly to the enzyme, the Kd's ranging from 3.1.10(-4) M with glycinol-AMP ester to 3.7.10(-9) M with L-isoleucinol-AMP ester. The binding is not affected by magnesium. It is shown that the free energies of binding of the esters can be calculated adding a constant contribution of the AMP-moiety of about - 4.1 (- 17) kcal/mole (kJ/mole) to the free energies of binding of the cognate amino acids, which we have reported earlier (19, 25, 26).
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PMID:On the binding of aminoalkyl adenylates to isoleucyl-tRNA synthetase from Escherichia coli MRE 600. 32 20

The inhibitors histidine and AMP cause the enzyme ATP phosphoribosyltransferase of E. coli to associate into a hexamer from its initial dimeric form. The behaviour of these inhibitors has been studied by three different methods. I) Equilibrium dialysis studies have shown that one mole of dimeric enzyme (67,000 g) binds one mole of histidine. II) By kinetic inhibition of the reaction studied at 21, 25 and 38 degrees C the enthalpy changes in the process of histidine and of AMP inhibition have been deduced. The inhibition has also been studied in function of enzyme concentration and temperature. The inhibition appears to be slightly negatively cooperative for histidine and positively cooperative for AMP. In neither case is it possible to obtain 100% maximal inhibition. III) By microcalorimetric analysis the values obtained for the enthalpies of histidine and of AMP interaction with the enzyme are similar.
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PMID:The action of inhibitors (histidine and AMP) on the ATP phosphoribosyltransferase of E. coli. 35 36

Antibodies to adenosine were elicited in rabbits by immunization with bovine serum albumin-adenosine conjugate. The antibodies were purified and fractionated on two affinity columns (Sepharose-oligo(A) and Sepharose-AMP). Two families of antibodies have been obtained. The antibodies purified on the Sepharose-oligo(A) column react with poly(A) while those purified on the Sepharose-AMP column do not, as shown by gel diffusion. The association constants for the binding of Fab fragments or IgG purified on the Sepharose-oligo(A) column and several haptens were deduced from dialysis equilibrium, fluorescence quenching and displacement of AMP-fluorescein conjugate. The antibodies mainly recognize adenine, and the ribose or the phosphate group of (or AMP derivatives) do not play a critical role in the interaction. Thermodynamic parameters for adenosine-Fab fragments complexes have been determined deltaH degrees = 16 kcal/mole and deltaS degrees = - 15 cal. degree-1 mole-1. Circular dichroism studies indicate that about three nucleotide residues penetrate the binding site of Fab fragments.
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PMID:Purification and specificity of antibodies to adenosine. 40 59

The interaction between phosphorylase B and an AMP analog, adenosine-5'-chloromethylphosphonate, is found to be irreversible. Their binding stechiometry is calculated from the differential absorption spectrum. Maximal inhibition is reached when 1,3--1,5 moles of the analogue is bound per mole of monomer phosphorylase B. The enzyme-inhibitory complex exhibited 50% activity is characterized by a sigmoid curve of the reaction rate dependency on the substrate concentration, by a decrease of the affinity to glucose-1-phosphate and the maximal rate, and by an increase of Hill's coefficient. Similar SH-groups titration curves were obtained for modified and native phosphorylase in the presence of AMP. Apophosphorylase was incapable of the complete reactivation by pyridoxalphosphate in the presence of adenosine-5'-chloromethylphosphonate. The complex of phosphorylase B and the AMP analogue is found to be inhibited by glucose-6-phosphate, like the native enzyme in the presence of AMP. The results of ultracentrifugation and disc electrophoresis show that the AMP analogue contributes the formation of the tetrameric form of the enzyme. The data obtained indicate the tight binding of adenosine-5'-chloromethylphosphonate in the active site of phosphorylase B. The properties of the complex confirm the importance of the phosphate group for the AMP binding in the allosteric site and for the enzyme activation.
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PMID:[Reaction between adenosine-5'-chloromethylphosphonate and the allosteric center of phosphorylase B]. 71 60

The effect of Mg2+ on the binding of adenylates to isolated chloroplast coupling factor 1 (CF1) was studied using CD spectrometry and ultrafiltration. At adenylate concentrations smaller than 100 muM, one mole of CF1 binds three moles of ATP (or ADP) regardless of the presence of Mg2+. In the presence of Mg2+, the first two ATP's bind to CF1 independently with the same binding constant of 2.5 X 10(-1) muM-1, then the third ATP binds with a much higher affinity of 10 muM-1. In the absence of Mg2+, the first ATP binds to CF1 with a binding constant of 2.5 X 10(-1) muM-1 then the other two ATP's bind less easily with the same binding constant of 4.0 X 10(-2) muM-1. The binding mode of ADP to CF1 is quite similar to that of ATP. In the presence of Mg2+, the binding constants of the first two ADP's are both 7.6 X 10(-2) muM-1, that of the third ADP being 4.0 muM-1. In the absence of Mg2+, the binding constant of the first ADP is 7.6 X 10(-2) muM-1, the constants of the other two ADP's both being 4.0 X 10(-2) muM-1. AMP caused a negligible change in CD.
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PMID:Magnesium ion-induced changes in the binding mode of adenylates to chloroplast coupling factor 1. 100 84

The pyruvate dehydrogenase complex from Axotobacter vinelandii was isolated in a five-step procedure. The minimum molecular weight of the pure complex is 600,000, as based on an FAD content of 1.6 nmol-mg protein-1. The molecular weight is 1.0-1.2 X 10(6), indicating 1 mole of lipoamide dehydrogenase dimer per complex molecule. Sodium dodecylsulphate gel electrophoretical patterns show that apart from pyruvate dehydrogenase (Mr89,000) and lipoamide dehydrogenase (Mrmonomer 56,000) two active transacetylase isoenzymes are present with molecular weight on the gel 82,000 and 59,000 but probably actually lower. The pure complex has a specific activity of the pyruvate-NAD+ reductase (overall) reaction of 10 units-mg protein-1 at 25 degrees C. The partial reactions have the following specific activities in units-mg protein-1 at 25 degrees C under standard conditions: pyruvate-K3Fe(CN)6 reductase 0.14, transacetylase 3.6 and lipoamide dehydrogenase 2.9. The properties of this complex are compared with those from other sources. NADPH reduced the FAD of lipoamide dehydrogenase as well in the complex as in the free form. NADP+ cannot be used as electron acceptor. Under aerobic conditios pyruvate oxidase reaction, dependent on Mg2+ and thiamine pyrophosphate, converts pyruvate into CO2 and acetate; V is 0.2 mumol 02-min-1-mg-1, Km(pyruvate)0.3 mM. The kinetics of this reaction shows a linear 1/velocity-1/[pyruvate] plot. K3Fe(CN)6 competes with the oxidase reaction. The oxidase activity is stimulated by AMP and sulphate and is inhibited by acetyl-CoA. The partially purified enzyme contains considerable phosphotransacetylase activity. The pure complex does not contain this activity. The physiological significance of this activity is discussed.
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PMID:The pyruvate-dehydrogenase complex from Azotobacter vinelandii. 120 21

Purified ribulose-bisphosphate carboxylase (EC 4.1.1.39) was strongly and equally inhibited either by ADP or GDP and to a lesser extent by IDP. AMP or ATP exerted little effect on activity. Inhibition by the nucleotide diphosphates was competitive with respect to RuBP and non-competitive with respect to "CO2" and Mg2+, respectively. Treatment of the enzyme with urea or guanidine-HCl resulted in rapid loss of activity that was not restored by dialysis even in the presence of Mg2+ and cysteine. Sodium dodecyl sulfate electrophoresis of 8.0 M urea treated enzyme revealed the presence of a fast-moving (small) sub-unit with molecular weight 14150 and a slower moving (large) sub-unit with molecular weight 68000. Examination of native enzyme by sodium dodecyl sulfate electrophoresis gave sub-units of 13700 and 55500 respectively. The amino acid content standardized to phenylalanine was essentially similar to that from other sources. Arrhenius plots showed a "break" at 29 degrees C with an Ea of 12.34 kcal per mole for the steeper part of the curve and a deltaH of 11.43 kcal per mole while for the less steep region, the Ea was 1.04 kcal per mole and the deltaH 1.92 kcal per mole.
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PMID:Physical properties and metabolite regulation of ribulose bisphosphate carboxylase from Thiobacillus A2. 127 53

A simple and sensitive enzymatic method for determination of plasma and serum fatty acids (FAs) is described. The method is based on acylation of long chain FAs by a bacterial acyl-CoA synthetase (ACS) producing equivalent amounts of acyl-CoA and AMP. AMP production was measured using the coupled reaction of myokinase (MK), pyruvate kinase (PK) and lactate dehydrogenase (LDH) allowing fluorinate detection of NADH. Two moles of NAD were produced per mole of FA acylated. Concentrations of substrates and enzymes were kept as low as possible maintaining the ACS reaction as rate limiting. Addition of fat-free human serum albumin (HSA) to standards reduced initial reaction rates but did not affect end-point fluorescence levels. Triton X-100 partly counteracted the inhibition by HSA. To keep albumin concentration low, plasma or serum samples were diluted by 1:400. Duplicate measurements of plasma or serum FA concentrations between 0 and 2 mmol l-1 can then be performed on 5 microliters samples with intra- and inter-assay variation coefficients of 1.7 and 4% respectively.
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PMID:Enzymatic microdetermination of plasma and serum free fatty acids. 145 65


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