Regulation of phosphoenolpyruvate carboxylase of Zea mays by metabolites

Crude preparations of phosphoenolpyruvate carboxylase obtained from aetiolated seedlings of Zea mays are unstable but can be stabilized with glycerol. At the pH optimum of 8.3, the Km value for phosphoenolpyruvate is 80μm. When assayed at 30°C, the enzyme shows normal Michaelis–Menten kinetics, but when assayed at 45°C sigmoid kinetics are exhibited. At pH7.0 the enzyme is inhibited by a number of dicarboxylic acids and by glutamate and aspartate. d and l forms of the hydroxy acids and amino acids are inhibitory and the kinetics approximate to simple non-competitive inhibition. The same compounds produce less inhibition at pH7.6 than at pH7.0 and the kinetics of inhibition are more complex. The enzyme is activated by Pi, by SO42- and by a number of sugar phosphates. Maximum activation occurs at acid pH values, where enzyme activity is lowest. The enzyme is activated by AMP and inhibited by ADP and ATP so that the response to energy charge is of the R type and is thus at variance with Atkinson's (1968) concept of energy charge. The physiological significance of the response to metabolites is discussed.

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The Kinetics of Inhibition of the Action of Thrombin by the Fibrinopeptides Formed during the Clotting of Fibrinogen

Thrombosis and Haemostasis ◽

10.1055/s-0038-1655562 ◽

1966 ◽

Vol 16 (01/02) ◽

pp. 277-295 ◽

Cited By ~ 1

Author(s):

A Silver ◽

M Murray

Keyword(s):

Amino Acids ◽

Competitive Inhibition ◽

Amorphous Material ◽

Peptide Bond ◽

Initial Reaction ◽

Reaction Products ◽

Coagulation Mechanism ◽

Kinetics Of ◽

Kinetics Of Inhibition ◽

Burk Plot

SummaryVarious investigators have separated the coagulation products formed when fibrinogen is clotted with thrombin and identified fibrinopeptides A and B. Two other peaks are observed in the chromatogram of the products of coagulation, but these have mostly been dismissed by other workers. They have been identified by us as amino acids, smaller peptides and amorphous material (37). We have re-chromatographed these peaks and identified several amino acids. In a closed system of fibrinogen and thrombin, the only reaction products should be fibrin and peptide A and peptide B. This reasoning has come about because thrombin has been reported to be specific for the glycyl-arginyl peptide bond. It is suggested that thrombin also breaks other peptide linkages and the Peptide A and Peptide B are attacked by thrombin to yield proteolytic products. Thrombin is therefore probably not specific for the glycyl-arginyl bond but will react on other linkages as well.If the aforementioned is correct then the fibrinopeptides A and B would cause an inhibition with the coagulation mechanism itself. We have shown that an inhibition does occur. We suggest that there is an autoinhibition to the clotting mechanism that might be a control mechanism in the human body.The experiment was designed for coagulation to occur under controlled conditions of temperature and time. Purified reactants were used. We assembled an apparatus to record visually the speed of the initial reaction, the rate of the reaction, and the density of the final clot formed after a specific time.The figures we derived made available to us data whereby we could calculate and plot the information to show the mechanism and suggest that such an inhibition does exist and also further suggest that it might be competitive.In order to prove true competitive inhibition it is necessary to fulfill the criteria of the Lineweaver-Burk plot. This has been done. We have also satisfied other criteria of Dixon (29) and Bergman (31) that suggest true competitive inhibition.

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Kinetics of phosphoenolpyruvate carboxylase from Zea mays leaves at high concentration of substrates

Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology ◽

10.1016/s0167-4838(01)00148-0 ◽

2001 ◽

Vol 1546 (1) ◽

pp. 242-252 ◽

Cited By ~ 6

Author(s):

Alejandro Tovar-Méndez ◽

Rosario A. Muñoz-Clares

Keyword(s):

Zea Mays ◽

Phosphoenolpyruvate Carboxylase ◽

High Concentration ◽

Kinetics Of

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Inhibition of plasmin by fibrinogen

Biochemical Journal ◽

10.1042/bj2690299 ◽

1990 ◽

Vol 269 (2) ◽

pp. 299-302 ◽

Cited By ~ 10

Author(s):

A A R Higazi ◽

M Mayer

Keyword(s):

Competitive Inhibition ◽

Hill Coefficient ◽

Coagulation System ◽

Amidolytic Activity ◽

Plasmin Activity ◽

Straight Line ◽

Non Linear ◽

The Hill ◽

Kinetics Of ◽

Kinetics Of Inhibition

The kinetics of inhibition of the amidolytic activity of plasmin on D-Val-L-Leu-L-Lys p-nitroanilide hydrochloride (S-2251) by fibrinogen and fibrin were determined. Reciprocal (1/v versus 1/[S]) plots of plasmin inhibition by 0.50 microM-fibrinogen showed a non-linear downward curve. The Hill coefficient (h) was 0.68, suggesting negative co-operativity. By contrast, fibrin produced a simple competitive inhibition of plasmin (Ki = 12 micrograms/ml). Addition of 0.1 mM-6-aminohexanoic acid shifted the non-linear curve obtained in the presence of fibrinogen to a straight line as for controls, indicating that 6-aminohexanoic acid abolishes the fibrinogen-induced inhibition. Transient exposure of the enzyme to pH 1.0 abrogates the ability of fibrinogen to inhibit plasmin activity. Acidification had no effect on the Vmax but increased the Km of plasmin. The present evidence for modulation of plasmin reveals a novel mechanism for control of fibrinolysis by fibrinogen, a component of the coagulation system and the precursor of the physiological substrate of plasmin.

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Adenine nucleotides and magnesium ions in relation to control of mammalian cerebral-cortex hexokinase

Biochemical Journal ◽

10.1042/bj1120579 ◽

1969 ◽

Vol 112 (5) ◽

pp. 579-586 ◽

Cited By ~ 40

Author(s):

H S Bachelard ◽

P. S. G. Goldfarb

Keyword(s):

Mixed Type ◽

Competitive Inhibition ◽

Adenine Nucleotides ◽

Effective Control ◽

Magnesium Ions ◽

Soluble Enzyme ◽

Kinetics Of ◽

Kinetics Of Inhibition

1. The kinetics of inhibition of brain soluble cytoplasmic hexokinase by ADP were examined in relation to variations in the concentrations of Mg2+ and ATP. The type of inhibition observed was dependent on the Mg2+/ATP ratio. 2. ADP at Mg2+/ATP ratios 2:1 exhibited inhibition of the ‘mixed’ type; at Mg2+/ATP ratios 1:1 the inhibition appeared to be competitive with regard to ATP. 3. Inhibition by free ATP was observed when the Mg2+/ATP ratio was less than 1:1. The inhibition was also of the ‘mixed’ type with respect to MgATP2−. 4. The inhibitions due to ADP and to free ATP were not additive. The results suggested that there may be up to four sites in the soluble enzyme: for glucose, glucose 6-phosphate, ADP and MgATP2−. 5. The ‘free’ non-particulate intracellular Mg2+ concentration was measured and concluded to be about 1·5mm. 6. The concentrations in vivo of Mg2+ and ATP likely to be accessible to a cytoplasmic enzyme are suggested to be below those that yield maximum hexokinase rates in vitro. The enzymic rates were measured at relevant suboptimum concentrations of Mg2+ and ATP in the presence of ADP. Calculations that included non-competitive inhibition due to glucose 6-phosphate (56–65% at 0·25mm) resulted in net rates very similar to the measured rates for overall glycolysis. This system may therefore provide a basis for effective control of cerebral hexokinase.

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Kinetics of Inhibition of ADP-Induced Platelet Aggregation by Dihomo-β-Linolenic Acid (DHLA).

10.1055/s-0039-1684703 ◽

1979 ◽

Author(s):

M. Zuzel ◽

A. Spencer

Keyword(s):

Fatty Acids ◽

Platelet Aggregation ◽

Competitive Inhibition ◽

Human Platelets ◽

Inhibition Of Platelet Aggregation ◽

Kinetics Of ◽

Dose Dependent ◽

Kinetics Of Inhibition ◽

Primary Platelet

In the study of the effects on platelets of thromboxanes and prostaglandins (PG) large amounts of precursor fatty acids have frequently been added to platelet suspensions. In the case of DHLA, this results in a general inhibition of platelet reactions. We have employed kinetic analysis of the inhibition of ADP-induced primary platelet aggregation to estimate the potency, specificity and mode of action of DHLA on human platelets in citrated PRP. Platelet PG production was estimated from formation of malondialdehyde (MDA) and aspirin (ASA) was used to inhibit production-Inhibition of aggregation and MEA formation were dose-dependent and both were observed at DHLA concentrations of 0.1 mM and above. Inhibition of aggregation was of mixed type, consisting of an ASA-sensitive competitive component (K1 ≈ 0.2mM) and an ASA-insensitve component which was non-competitive and dominated inhibition at higher concentrations of DHLA. The KI (DHLA) of the non-competitive component varied from 0.4 to 1.5 mM with different batches of PRP. Other polyunsaturated fatty acids which are not PG precursors, did not cause competitive inhibition but were as potent as DHLA in the non-competitive inhibition of aggregation. The results show that a large part of the inhibition of platelet aggregation by DHLA in vitro is not due to its transformation into an inhibitor of platelet function in the PG production pathway.

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Purification and kinetics of phenylpropanoid O-methyltransferase activities from Brassica oleracea

Biochemistry and Cell Biology ◽

10.1139/o89-115 ◽

1989 ◽

Vol 67 (11-12) ◽

pp. 763-769 ◽

Cited By ~ 14

Author(s):

Emidio De Carolis ◽

Ragai K. Ibrahim

Keyword(s):

Competitive Inhibition ◽

Divalent Cations ◽

Product Inhibition ◽

Affinity Column ◽

Ph Optimum ◽

Substrate Interaction ◽

Interaction Kinetics ◽

Molecular Masses ◽

Inhibition Studies ◽

Kinetics Of

Two phenylpropanoid O-methyltransferase isoforms were purified to homogeneity from young cabbage leaves. They catalyzed the meta-O-methylation of caffeic and 5-hydroxyferulic acids to ferulic and sinapic acids, respectively. Both isoforms I and II exhibited different elution patterns from a Mono Q column, distinct apparent pIs on chromatofocusing, different product ratios, and stability on adenosine–agarose affinity column. On the other hand, both isoforms had similar apparent molecular masses (42 kilodaltons) and a pH optimum of 7.6. They exhibited no requirement for divalent cations and were both irreversibly inhibited by iodoacetate. Substrate interaction kinetics of the more stable isoform I, using the 5-hydroxyferulic acid and S-adenosyl-L-methionine, gave converging lines. Product inhibition studies showed competitive inhibition between S-adenosyl-L-methionine and S-adenosyl-L-homocysteine and non-competitive inhibition between the phenylpropanoid substrate and its methylated product. The kinetic patterns are consistent with an ordered bi bi mechanism, where S-adenosyl-L-methionine is the first substrate to bind and S-adenosyl-L-homocysteine is the last product released.Key words: phenylpropanoid O-methyltransferase, purification, isoforms, adenosine–agarose affinity chromatography, kinectic mechanism.

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Intact mesophyll protoplasts from Zea mays as a source of phosphoenolpyruvate carboxylase unaffected by extraction: Advantages and limitations

Physiologia Plantarum ◽

10.1034/j.1399-3054.1990.800417.x ◽

1990 ◽

Vol 80 (4) ◽

pp. 605-611 ◽

Cited By ~ 1

Author(s):

Y. Petropoulou ◽

Y. Manetas ◽

N. A. Gavalas

Keyword(s):

Zea Mays ◽

Phosphoenolpyruvate Carboxylase ◽

Mesophyll Protoplasts

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Kinetic Aspects of the Interaction of Blood Clotting Enzymes

Thrombosis and Haemostasis ◽

10.1055/s-0038-1651214 ◽

1968 ◽

Vol 19 (03/04) ◽

pp. 364-367 ◽

Cited By ~ 4

Author(s):

H. C Hemker ◽

P. W Hemker

Keyword(s):

Enzyme Kinetics ◽

Blood Clotting ◽

Competitive Inhibition ◽

Competitive Inhibitor ◽

Kinetics Of

SummaryThe enzyme kinetics of competitive inhibition under conditions prevailing in clotting tests are developed and a method is given to measure relative amounts of a competitive inhibitor by means of the t — D plot.

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