The adenosine triphosphate inhibition of the pyruvate kinase reaction and its dependence on the total magnesium ion concentration

1. The effects of ATP, PP(i) and EDTA on the skeletal-muscle pyruvate kinase reaction at various concentrations of magnesium (where ;magnesium' refers to total Mg(2+), both free and in the form of complexes) were investigated. The reaction rate was determined as the amount of pyruvate formed in a recorded time of incubation. 2. At 44mm-magnesium the K(m) values for ADP and phosphoenolpyruvate were unaltered by the presence of ATP up to 6.8mm in systems buffered with either tris-hydrochloric acid or glycylglycine-sodium hydroxide, but the K(m) values were different in these systems. The K(m) for one substrate was independent of the concentration of the second substrate. 3. At 10mm-magnesium in the tris-hydrochloric acid system ATP inhibited the reaction competitively with respect to ADP and phosphoenolpyruvate. In the glycylglycine-sodium hydroxide system the inhibition appeared to be non-competitive. At 10mm-magnesium the K(m) values were lower than at 44mm-magnesium and dependent on the system used. 4. In the tris-hydrochloric acid system the reaction rate rose with increasing magnesium concentration up to a maximum at a concentration 10-20 times that of ADP. Further increase inhibited the reaction and at 44mm-magnesium the rate was 25-50% of its maximum. This inhibition paralleled that produced by increasing trimethylammonium chloride concentrations and was not due to a specific effect of the Mg(2+) ion. 5. In the presence of 6.8mm-ATP no reaction occurred below 4-6mm-magnesium, and further increase apparently abolished the inhibition as the reaction rate increased and became equal to those obtained in the absence of ATP at 10-25mm-magnesium. Further increase in magnesium concentration gave reaction rates that were slightly higher in the presence of ATP than in its absence. The maximal rate in the presence of ATP was distinctly lower than in its absence. When 6.8mm-PP(i) or 6.8mm-EDTA was present the variations in reaction rate with rising magnesium concentration were similar to that obtained in the presence of ATP below 6-8mm-magnesium but further increase in the magnesium concentration resulted in an increase in the rate up to a maximum comparable with that of the control. The effect of pure chelation was thus a displacement of the reaction maximum to higher magnesium concentrations without changing the maximal rate. When correction had been made for this effect, ATP gave inhibition at 44mm-magnesium that was competitive with respect to ADP (K(i) 2.1x10(-2)m). This degree of inhibition is far less than was reported earlier and its importance for the mechanism of the pyruvate kinase reaction is discussed.     

Synthesis of 10,11-dihydrofarnesyl pyrophosphate from 6,7-dihydrogeranyl pyrophosphate by prenyltransferase

The syntheses of 6,7-dihydrogeraniol and of its pyrophosphate are described. It is shown that this analogue of geranyl pyrophosphate is a substrate for liver prenyltransferase and that the product synthesized by this enzyme from it and isopentenyl pyrophosphate is 10,11-dihydrofarnesyl pyrophosphate. The K(m) value for 6,7-dihydrogeranyl pyrophosphate was determined to be 1.11+/-0.19mum as compared with 4.34+/-1.71mum for geranyl pyrophosphate. The maximum reaction velocity with the artifical substrate was, however, only about one-fourth of that observed with geranyl pyrophosphate. The binding of isopentenyl pyrophosphate to the enzyme was not affected by the artificial substrate.     

Analogues of geranyl pyrophosphate as inhibitors of prenyltransferase

Six analogues of geranyl pyrophosphate (the monophosphates of geraniol and tetrahydrogeraniol, and the pyrophosphates of nerol, octan-1-ol, tetrahydrogeraniol and citronellol) were synthesized, and were found to be inhibitors of pig liver prenyl- (geranyl-)transferase. The effects of each analogue were analysed in kinetic experiments, which showed the pyrophosphates of citronellol, tetrahydrogeraniol and octan-1-ol to be the most potent inhibitors. The results are interpreted to support a previous hypothesis that the main forces in the binding of substrates to prenyltransferase are non-specific lipophilic forces and a pyrophosphate-binding force.     

Transformation in Rhizobium japonicum

Summary The transformation of streptomycin resistance in Rhizobium japonicum was studied. The susceptible strain 211 was selected from sixty strains and one step mutant resistant to streptomycin in concentration 1 mg per 1 ml was used as the donor. The peak of the competence curve appeared at the ninth hour of growth; the frequency, when the homologous strain had been used was 0.01 p.c. The transformed resistance was of the same level as in the donor strain.This investigation forms part of a contribution prepared by the Czechoslovak National Committee for the International Biological Programme (Section PP: Production Processes).     

Isoamylases in blood donors

Summary Sera of 1,000 blood donors were tested for various combinations of salivary and pancreatic amylase isoenzymes and the frequency of their occurrence was determined in the series mentioned. Five combinations of isoamylases were found. A combination of 1 salivary and 1 pancreatic amylase was found most frequently (89.5%), the frequency of the other four combinations was relatively low (0.2–5.1%).Hereditary character of amylase isoenzymes was confirmed in a series of 36 families.     

On the validity of the genusDactylococcopsis (Cyanophyceae)

Summary The genusDactylococcopsis Hansg. 1888 (Cyanophyceae) is based on the typeD. rupestris, which was later identified as a green algae. Most of the many species described later were also placed to other groups of algae. Several authors even doubted about the existence of the genus. As, however, some species of Cyanophyceae correspond to the original generic diagnosis, the name Dactylococcopsis Hansg. ex R. et F.Chod. 1925 has been proposed as a nomen conservandum, and a new type (D. smithii R. et F.Chod.) has been defined. Further speciesD. linearis Geitl. 1935 and D.Planctonica Teil. 1942 has been unambiguously described till now.     

Zur Populationsgenetik der sauren Phosphatase der Erythrocyten (EC: 3.1.3.1): Phänotypen- und Allelhäufigkeiten in der ČSSR

Summary Within a population sample of 307 blood donors of Prague the polymorphism of the red cell acid phosphatase has been investigated. Gene frequency estimates are: P^a=0.365, P^b=0.578, P^c=0.057.     

Der Einfluß einiger Hormone auf die heteroplastische Knorpel- und Knochenbildung im Herzmuskel der Ratte

Zusammenfassung Es wird über histopathologische Untersuchungen des chronologischen Verlaufs an der durchSelyes Ventrikel-Ligatur hervorgerufenen Herzspitzennekrose bei Ratten berichtet. In dem der Nekrose verfallenen Myokard setzt alsbald Organisation ein; dann wandelt sich allmählich das Granulationsgewebe in eine zellarme und faserreiche Narbe um. In den subendokardialen Schichten des Narbengewebes entstehen erst Knorpel-, später Knochenherde. Der Prozeß der Knorpelbildung wird nicht von Verkalkung eingeleitet; vielmehr werden zuerst die Fibroblasten zu Knorpelzellen, um die sich dann sekundär Kalziumsalze ablagern; danach spielt sich eine typische endochondrale Ossifikation ab. Schließlich erscheint Knochenmarkgewebe zwischen den Knochenbalken.Triamcinolon hemmt geringgradig die Bindegewebsproliferation, Thyroxin steigert die Knorpel- und Knochenbildung, während Östradiol diese Vorgänge nicht beeinflußt.

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The effect of hormones on the heteroplastic cartilage and bone formation in the cardiac muscle of the rat

Summary A chronologic study was made of the histopathologic changes which occur in the cardiac apex of the rat following the application of Selye's ventricular ligature. Organisation in the necrotic cardiac muscle begins soon after ligature. Later, the granulationtissue is gradually replaced by scar tissue which is poor in cells but rich in fibers. In the subendocardial fibrous tissue, cartilage and bone develop. It is emphasized that cartilage formation is not initiated by calcification. Instead, the fibroblasts are converted to cartilage cells and, later, calcium salts are deposited in the matrix. This is followed by endochondral bone formation. Finally, bone marrow appears in the intertrabecular spaces.Triamcinolone mildly hindered connective-tissue proliferation, thyroxine increased cartilage and bone formation, while estradiol did not influence these processes.

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Die Versuche, die diesem Bericht zugrunde liegen, wurden durch das Ministère de la Santé, Québec, die Quebec Heart Foundation, Montreal, das Medical Research Couneil of Canada (Block Term Grant MT-1829) und das USPHS, Child Welfare (Grant HDO 2612-02) unterstützt. Die Autoren danken an dieser Stelle der Firma Lederle Laboratories Div., Pearl River, N.Y., USA für das bei diesen Versuchen verwendete Triamcinolon (Aristocort^®) und der Firma Schering Corporation Ltd., Pointe Ciaire, Quebec, für die Bereitstellung von Estradiol.