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1.
《The International journal of biochemistry》1991,23(10):991-995
- 1.1. Purified ostrich (Struthio camelus) liver fructose-1,6-bisphosphatase exhibited an absolute requirement for Mg2+.
- 2.2. The enzyme catalyzed the hydrolysis of fructose-1,6-bisphosphate, sedoheptulose-l,7-bisphosphate and ribulose-l,5-bisphosphate.
- 3.3. S0.5 for substrate was 1.4 μM.
- 4.4. AMP was a potent non-competitive inhibitor with respect to substrate (Ki of 25 μM).
- 5.5. Fructose-2,6-bisphosphate was a potent competitive inhibitor of the enzyme (Ki of 4.8 μM).
2.
《The International journal of biochemistry》1989,21(12):1337-1342
- 1.1. A novel glycogen phosphorylase inhibitor was partially purified from crayfish hepatopancreas.
- 2.2. The inhibitor was found only in two species of crayfish examined, and not in lobster, fresh and salt water clams, mussels or cockroaches.
- 3.3. The inhibitor is a small protein (Mr = 23,000) which did not show proteolytic activity.
- 4.4. Preliminary kinetic analysis of the inhibitory mechanism indicated that it bound to both glycogen and the glycogen phosphorylase protein.
- 5.5. Inhibitor binding to glycogen resulted in a competitive inhibition pattern with respect to glycogen phosphorylase (inhibition constant of ca 10 μg/ml).
- 6.6. The inhibitor also bound glycogen phosphorylase directly with a binding coefficient of 100 μg/ml resulting in a partially non-competitive inhibition pattern with respect to phosphate.
3.
《The International journal of biochemistry》1980,11(1-2):145-149
- 1.1. Glutamate dehydrogenase flux by rat kidney mitochondria incubated with 1 mM glutamine plus 2–3 mM glutamate was stimulated by aminooxyacetate. This effect was inhibited by α-ketoglutarate.
- 2.2. Studies with intact mitochondria and mitochondrial sonicates revealed a linear inverse relationship between glutamate deamination and α-ketoglutarate levels.
- 3.3. The data revealed that α-ketoglutarate is a competitive inhibitor of glutamate dehydrogenase with an apparent Ki of 0.6mM.
- 4.4. The data suggest that aminooxyacetate stimulates glutamate deamination by a mechanism mediated by α-ketoglutarate.
4.
《The International journal of biochemistry》1993,25(8):1157-1164
- 1.1. A proteinaceous inhibitor for S-adenosyl-l-methionine (AdoMet)-dependent transmethylation reactions has been purified to apparent homogeneity from rat liver cytosolic fraction.
- 2.2. The peptide was made up of 29 amino acid residues with a molecular weight of 2,584. Glycine accounted for 52% of the total amino acids.
- 3.3. Employing AdoMet: protein-carboxyl O-methyltransferase (Protein methylase II) and bovine serum γ-globulin as in vitro substrate, the mode of inhibition was found to be non-competitive with Ki value of 1.9 × 10−8 M.
- 4.4. When the inhibitor was present in the reaction mixture together with S-adenosyl-l-homocysteine (AdoHcy), which is a competitive inhibitor for AdoMet, the extent of inhibition exceeded that exerted by each individual inhibitor alone, suggesting that the sites of the inhibitors on the enzyme molecule are different.
- 5.5. Almost a stoichiometric relationship exists between the enzyme and the inhibitor molecule, the ratio being approx one.
5.
《Comparative biochemistry and physiology. B, Comparative biochemistry》1993,104(4):925-932
- 1.1. In order to assign a meaningful role to the phosphorolytic pathway in Mytilus glycogen metabolism the kinetic mechanism of phosphorylase b, and its allosteric control, were studied.
- 2.2. The kinetic parameters of phosphorylase b from the mussel Mytilus galloprovincialis were determined. Michaelis constants (Km or S0.5) were in the range of 0.32–2.49 mg/ml for glycogen, 7–16 mM for Pi and 114–423 μM for AMP. In the direction of glycogen synthesis, the Km value for glucose-1-P was approximately 180 mM.
- 3.3. The enzyme displayed homotropic co-operativity towards the binding of co-substrate and AMP (Hill coefficients of 2 and 1.4, respectively) and heterotropic co-operativity between substrates and AMP.
- 4.4. The concentration of glycogen in the Mytilus mantle is between 38- and 125-fold higher than the apparent Km of phosphorylase b; the concentration of AMP varies throughout the year from 10 to 175 μM, up to a value close to the apparent Km for the effector.
- 5.5. The apparent Km for Pi is close to the concentration found in the mantle. This ligand showed more important regulatory effects than the effector AMP.
6.
《The International journal of biochemistry》1992,24(7):1017-1021
- 1.1. The properties of Na+/K+-transporting ATPase in microsomal fractions from the nervous tissue of the grasshopper, Poekilocerus bufonius were investigated.
- 2.2. Two components of ATPase activity are present.
- 3.3. Inclusion of 1 mM ouabain in the incubation media reduced the activity of total and Na+/K+-ATPase by 57 and 79%, respectively.
- 4.4. The maximum velocity (Vmax) was decreased by the addition of 1 mM ouabain, whereas the apparent Km value was not affected indicating a non-competitive type of inhibition.
- 5.5. The calculated value of the pI50 was 6.4 (I50 = 3.98 × 10−7M) for ouabain inhibition of the enzyme showing great sensitivity to the cardiac glycoside ouabain.
- 6.6. The present results show that the physicochemical properties of Na+/K+-transporting ATPase from the brain of P. bufonius are essentially the same as for the enzyme prepared from the excretory system of the insect which has been previously investigated.
- 7.7. Dissimilarities were also observed between these tissues in the way that the enzyme from the brain was sensitive to ouabain inhibition with a non-competitive type rather than a ouabain-resistance and a competitive type of inhibition for the enzyme from the excretory system.
- 8.8. These dissimilarities are probably due to different isoenzyme patterns available in the same insect.
7.
《The International journal of biochemistry》1989,21(7):783-789
- 1.1. Kinetic constant values of the reaction catalyzed by bass liver glucose 6-phosphate dehydrogenase show to be modified between 10 and 40°C.
- 2.2. The Arrhenius plot between 10 and 50°C shows two slopes with different activation energies.
- 3.3. These results suggest a regulation of this enzyme by environmental temperature.
- 4.4. Kinetics of ATP inhibition were examined between pH 6.2 and 7.8: patterns and Ki values obtained are affected by the pH variation.
- 5.5. NADH is an effective inhibitor of bass glucose 6-phosphate dehydrogenase but this enzyme does not show NAD-linked activity.
- 6.6. Kinetics of pyridoxal 5′-phosphate inhibition have indicated the presence of a lysine in the catalytic site for NADP+.
8.
《The International journal of biochemistry》1994,26(2):155-162
- 1.1. Pseudomonas aeruginosa phospholipase C from culture supernatants of bacteria grown in high-Pi basal salt medium with choline, as the sole carbon and nitrogen source, was purified by precipitation with 70% saturation ammonium sulfate in the presence of celite.
- 2.2. The PLC activity was eluted of this mixture by the use of a reverse gradient of 70-0% ammonium sulfate.
- 3.3. The peak containing the PLC activity revealed a single protein after SDS-PAGE.
- 4.4. The method could also be applied to purify PLC produced in a low-Pi complex medium. The resultant preparation was not homogeneous.
- 5.5. The molecular weight for both PLC preparations was about 70 kDa.
- 6.6. Both PLC used phosphatydilcholine and sphingomyelin as substrates, displayed hemolytic activity an exhibited an apparent KM of 25 mM for p-nitrophenylphosphorylcholine.
- 7.7. They were not inhibited by 1% sodium deoxycholate but were 30% inhibited by 1% Triton X-100.
- 8.8. 2% sodium dodecylsulfate and 1% tetradecyltrimethylammonium bromide inhibited the PLC from the HPl-BSM plus choline but not the enzyme from the LPl-CM.
9.
《The International journal of biochemistry》1976,7(1-2):67-71
- 1.1. Kinetic properties of the inhibitory effect of inorganic phosphate and fluoride and of the activatory effect of potassium ion on human skeletal muscle AMP-deaminase (E.C. 3.5.4.6) have been investigated
- 2.2. It has been shown that phosphate is a competitive inhibitor (K1, ≈0.8 × 10−3M) and fluoride a noncompetitive inhibitor (K1≈3.2 × 10−3 M) of human muscle AMP-deaminase.
- 3.3. The changes of potassium ion concentration between 20 and 200 mM did not influence the Michaelis constant which was about 0.9 x 10−3 M at 30°C. Also the change of substrate concentration in the range 40–300 μM did not influence the activation constant for potassium (Ka≈0.4 × 10−1 M).
- 4.4. Higher concentraion of potassium (200mM) was found to diminish the “temperature sensitivity” of the enzyme activity.
- 5.5. The energy of activation (E) in the presence of 150 mM KC1 calculated from Arrhenius plot was about 4600 cal/mole of substrate. The heat of the enzyme-substrate complex formation obtained from the plot of log Km vs T−1 was shown to have positive value (+2200 cal/mole) at the temperatures lower than 23°C and negative value (—4100 cal/mole) at the temperatures higher than 23°C.
10.
《The International journal of biochemistry》1992,24(10):1657-1660
- 1.1. A thermostable orthophosphoric monoester phosphohydrolase (EC 3.1.3.1) from Thermus sp strain Rt41A has been purified 400-fold to give a specific activity of 25 U/mg at 60°C in IM diethanolamine (pH 11.1).
- 2.2. The enzyme has a Mr of 160,000 and is trimeric.
- 3.3. The half-life of the enzyme is 5 min at 85°C.
- 4.4. The enzyme has a wide specificity for a number of phosphate monoesters.
- 5.5. The Hm of the enzyme is pH dependent, so the pH optimum of the enzyme is affected by the substrate concentration.
- 6.6. The enzyme is inhibited 50% by 20 mM Ca2+ or Mg2+.
- 7.7. The Ki for phosphate, EDTA-di sodium salt and arsenate (in 1 M diethanolamine, pH 11.1) is approx 1.2, 1.6 and 4mM respectively.
- 8.8. Urea (200 mM) is not inhibitory.
11.
《The International journal of biochemistry》1989,21(3):291-295
- 1.1. Covalent coupling of fluorescein to methotrexate (MTX) by a 5-carbon spacer yields a dihydrofolate reductase (DHFR) inhibitor (FMTX) with Ki = 11 nM.
- 2.2. FMTX shows a fluorescence quenching with respect to fluorescein which is relieved by binding to the enzyme.
- 3.3. The dissociation constants (Kd) of MTX, FMTX, NADPH and 7,8-dihydrofolate (DHF) from bovine liver DHFR have been determined by fluorometric titrations.
- 4.4. The Kd values for NADPH, MTX and FMTX from the complementary binary complexes (MTX·DHFR, FMTX·DHFR and NADPH·DHFR) were also obtained; these show a 2- to 4-fold decrease with respect to those obtained by titration of the free enzyme.
- 5.5. A competitive assay for MTX has been developed by exploiting the fluorescence enhancement of DHFR-bound FMTX. This assay may be useful for the routine determination of MTX in the concentration range from 10−9 to 10−7 M.
12.
《The International journal of biochemistry》1982,14(6):519-528
- 1.1. The non-specific hen's egg yolk alkaline phosphatase is a metalloprotein (Zn2+?) composed of two identical inactive subunits.
- 2.2. A second metal site preferably binds Mg2+ (15-fold activation). Me(II))H2O)H+, a charged arginine, and tyrosine in the active site are involved in positioning and binding of the substrate and metal ion.
- 3.3. Substrate inhibition differs with pH. This may be related to the presence of two active sites in the enzyme, one in each subunit.
- 4.4. Uncompetitive inhibition with L-phenylalanine and analogues suggests a phosphorylated intermediate.
- 5.5. Inhibition is weakly competitive with Pi strong non-competitive with PPi as compared to Mg2+-free PPi, and partially competitive with arsenate.
- 6.6. The purified enzyme is stabilized and activated by amines and proteins.
13.
《The International journal of biochemistry》1985,17(2):253-257
- 1.1. Pyruvate kinase from mycelium of Phycomyces blakesleeanus NRRL 1555(−) has been partially purified and some kinetic properties has been investigated at pH 7.5.
- 2.2. Positive homotropic interactions were observed with phosphoenolpyruvate and Mg2+, showing Hill coefficient values of 2.8 and 2.5, respectively, whereas hyperbolic kinetics are found when ADP was the variable substrate.
- 3.3. Fructose 1,6-bisphosphate acts as a heterotropic allosteric activator, markedly decreasing the S0.5 value for phosphoenolpyruvate saturation curve from a sigmoidal to a hyperbolic form.
- 4.4. ATP inhibits pyruvate kinase from mycelium of Phycomyces blakesleeanus. ATP appears to be a non-competitive inhibitor with respect PEP and competitive inhibitor with respect ADP.
14.
《Comparative biochemistry and physiology. A, Comparative physiology》1986,83(3):489-493
- 1.1. Fundamental chitin digestion characteristics of Crassostrea virginica crystalline style were investigated.
- 2.2. Optimum temperature and pH were 34°C and 4.8. respectively.
- 3.3. The colloidal regenerated chitin (0.56mol/0.5 ml: GlcNAc equivalents) was saturating under all enzyme levels encountered.
- 4.4. There was no evidence of end product inhibition, even after 100 hr incubation.
- 5.5. Calculated Km for the chitinase complex was 1.19mM when determined using a 30 min assay, but was only 0.70 mM when determined using a 4.6 hr assay.
- 6.6. Both Km values are lower than reported for similar assays in other molluscs and for most bacteria.
- 7.7. Effect of substrate preparation on the kinetics are discussed.
- 8.8. Eight peaks of chitinase activity were resolved by DEAE-Fractogel ion exchange chromatography.
15.
《The International journal of biochemistry》1994,26(2):195-200
- 1.1. Glucose-6-phosphate dehydrogenase (G6PDH EC 1.1.1.49) from mouse liver has been purified 1100-fold by extraction, ion-exchange chromatography on DE-52, absorption chromatography on Bio-Gel HTP and gel filtration through sepharose 6 HR 10/30. The purified enzyme showed a single band in silver stained SDS-PAGE.
- 2.2. The native and subunit molecular weight were 117 and 31 kDa respectively.
- 3.3. The kinetic studies and the patterns obtained from the inhibition by-products suggest that the enzyme follows an ordered sequential kinetic mechanism.
- 4.4. The reduced Km values for the substrates favour the operativity of the enzyme. The “fine control” of the enzymatic activity was exerted by the NADPH, whose Ki is several fold lower than the in vivo concentration.
16.
《The International journal of biochemistry》1994,26(5):667-677
- 1.1. Highly purified 200 kDa casein kinase II from rabbit lactating mammary gland (MG-CK II) was obtained by means of a new purification procedure consisting of one phosphocellulose and three Mono Q steps.
- 2.2. Its Km for ATP was 2.22 μM and 0.57 mg/ml and 0.13 mg/ml for partially dephosphorylated casein and phosvitin respectively. Stathmine was also suitable as substrate. 2-aminopurine and 6-dimethylaminopurine inhibited efficiently MG-CK II Ki = 5 and 1 mM respectively).
- 3.3. MG-CK II autophosphorylated on its α-, α '- and β-subunits. The β-subunit auto-phosporylation was enhanced in presence of exogenous substrate. Its modulation was highly dependent on ATP concentration.
- 4.4. The effects of basic compounds which affected dramatically the phosphorylation of dephosphorylated casein in presence of various ATP concentrations were reported.
17.
《The International journal of biochemistry》1988,20(4):463-470
- 1.1. Partially purified rat liver ornithine decarboxylase is inhibited by several diamines including putrescine, 1,3-diaminopropane, cadaverine and p-phenylenediamine.
- 2.2. The inhibition is dependent on pH, being strong at pH above 8 and negligible below pH 6.5.
- 3.3. The kinetic study of the inhibition showed that while the aromatic diamine behaved as a simple competitive inhibitor, the aliphatic diamines presented a more complex pattern of inhibition in which two molecules of inhibitor might bind to the enzyme active site.
- 4.4. The KI values for the different inhibitors were calculated and the degree of affinity for the enzyme was p-phenylenediamine > putrescine > cadaverine > 1,3-diaminopropane.
- 5.5. A molecular mechanism explaining how one or two molecules of inhibitor can bind to the enzyme is proposed.
18.
《The International journal of biochemistry》1982,14(4):277-284
- 1.1. Glycogen, caffeine and glucose dissociate phosphorylase a tetramer to dimers with half-maximum effect at 0.16%, 1.1 and 71 mM concentration, respectively, as monitored by crosslinking with dimethyl suberimidate at 18°C.
- 2.2. The above ligands increase the rate of dephosphorylation and tryptic digestion of phosphorylase a at 18°C in the same way with half-maximum effect at 0.04%, 0.1 and 9 mM concentration, respectively.
- 3.3. Caffeine and glucose acted synergistically in tetramer dissociation as well as in the enzymic modifications.
- 4.4. The α-anomer of d-glucose was twice as effective as its mutarotational equilibrium solution.
19.
《The International journal of biochemistry》1985,17(10):1085-1089
- 1.1. Hydroxypyruvate reductase has been purified 193-fold from Lemna minor L. by affinity chromatography on Blue Sepharose.
- 2.2. The enzyme has activity over a broad pH range (optimum pH 6), a Km hydroxypyruvate of 59 μ M and Km NADH of 12μM.
- 3.3. Crude extracts of Lemna exhibit substrate inhibition of activity above 1 mM hydroxypyruvate, a property which is lost on purification.
- 4.4. Oxaloacetate inhibits purified preparations of the enzyme and a possible role for such regulation in vivo is discussed.
20.
《The International journal of biochemistry》1985,17(3):341-345
- 1.1. A new tetralysine endopeptidase from Escherichia coli AJ005 has been purified about 135-fold.
- 2.2. The peptidase seems to be specific to tetralysine among lysine homopolymers.
- 3.3. The optimal pH was about 7.5
- 4.4. The activity was inhibited by KCN but not inhibited by soybean trypsin inhibitor.
- 5.5. The apparent Km value was 2.5 × 1O−3 M for tetralysine.