Oyster citrate synthase: control of carbon entry into the Krebs cycle of a facultative anaerobe

1976 ◽  
Vol 54 (6) ◽  
pp. 892-895 ◽  
Author(s):  
J. H. A. Fields ◽  
P. W. Hochachka
Keyword(s):  
Enzyme Activity ◽  
Catalytic Activity ◽  
Crassostrea Gigas ◽  
Citrate Synthase ◽  
Specific Activity ◽  
Krebs Cycle ◽  
Adductor Muscle ◽  
Facultative Anaerobe ◽  
Wet Weight ◽  
Regulatory Properties

Citrate synthase (EC 4.1.3.7) in adductor muscle of the oyster, Crassostrea gigas, occurred in relatively low specific activity, about 1.5 μmol product formed per minute per gram wet weight of tissue. The enzyme activity was essentially independent of pH between pH 7.5 and 9.0. The Km values for acetyl-CoA and oxaloacetate were about 0.005 mM in each case. Catalytic activity was modulated by the adenylates, citrate, and 2-ketoglutarate, all of which were inhibitory. The regulatory properties of the enzyme suggest that during the transition to anoxia oxaloacetate becomes limiting, thus reducing flux through the initial stages of the Krebs cycle.

Moving into Nanotechnology Roles to Mimic and Boost Enzyme Activity

2018 ◽  
pp. 421-440
Author(s):  
Maria Laura Soriano
Keyword(s):  
Enzyme Activity ◽  
Catalytic Activity ◽  
Enzyme Immobilization ◽  
Mesoporous Materials ◽  
Specific Activity ◽  
Artificial Enzymes ◽  
Catalytic Nanoparticles ◽  
Solid Liquid ◽  
By Products ◽  
Insight Into

A new tendency toward the design of artificial enzymes based on nanostructures (nanodots, nanofibers, mesoporous materials) has emerged. On one hand, nanotechnology bestows self-catalytic nanoparticles with a specific activity to achieve efficient reactions with low number of by-products. On other hand, the nanoparticles may behave as nanometric scaffolds for hosting enzymes, promoting their catalytic activity and stability. In this case, enzyme immobilization requires the preservation of the catalytic activity by preventing enzyme unfolding and avoiding its aggregation. These approaches render many other advantages like hosting/storing enzymes in nanotechnological solid, liquid, and gel-like media. This chapter focuses on the most up-to-date approaches to manipulate or mimic enzyme activity based on nanotechnology, and offers examples of their applications in the most promising fields. It also gives new insight into the creation of reusable nanotechnological tools for enzyme storage.

Octopus mantle citrate synthase

1976 ◽  
Vol 54 (6) ◽  
pp. 886-891 ◽  
Author(s):  
J. H. A. Fields ◽  
H. Guderley ◽  
K. B. Storey ◽  
P. W. Hochachka
Keyword(s):  
Citrate Synthase ◽  
Specific Activity ◽  
Energy Charge ◽  
Krebs Cycle ◽  
Fold Increase ◽  
Adenylate Energy Charge ◽  
Michaelis Constant ◽  
Acetyl Coa ◽  
Muscle Work ◽  
Krebs Cycle Intermediates

Citrate synthase (EC 4.1.3.7) in mantle muscle of the octopus, Octopus cyanea, occurs in relatively low specific activity and is largely independent of pH between 7.5 and 9.0. Catalytic activity is regulated by the adenylate energy charge and by at least two Krebs cycle intermediates, α-ketoglutarate and citrate. Of the adenylates, ATP is by far the most potent inhibitor, at near-physiological concentrations (4 mM), causing almost a 20-fold increase in the Michaelis constant for acetyl-CoA. Citrate and α-ketoglutarate, on the other hand, are competitive with respect to oxaloacetate, rather than acetyl-CoA, and bring about large increases in the Michaelis constant for oxaloacetate. The regulatory properties of citrate synthase allow a curtailment of carbon flow into the Krebs cycle during periods of burst muscle work, when mantle anaerobic glycolysis is strongly activated.

On the role of octopine dehydrogenase in cephalopod mantle muscle metabolism

1976 ◽  
Vol 54 (6) ◽  
pp. 871-878 ◽  
Author(s):  
Jeremy H. A. Fields ◽  
John Baldwin ◽  
Peter W. Hochachka
Keyword(s):  
Enzyme Activity ◽  
Muscle Activity ◽  
Specific Activity ◽  
Ph Dependence ◽  
Muscle Metabolism ◽  
Ph Optimum ◽  
Wet Weight ◽  
Octopine Dehydrogenase

Octopine dehydrogenases from the mantle muscle of the squid, Symplectoteuthis oualaniensis, and of the octopus, Octopus ornatus, were kinetically characterized and compared. In the squid, the specific activity of the enzyme was about 110 μmol product formed per minute per gram wet weight; in the octopus that value was over 600. Both enzymes show similar pH dependence; in the direction of octopine formation the pH optimum was about 6.5, whereas in the direction of octopine oxidation it was about 8.5. The affinities for NADH, arginine, and pyruvate were similar (Km values were about 0.04 mM, 7 mM, and 2 mM respectively). Increasing the concentration of either arginine or pyruvate increased the affinity for the cosubstrate (pyruvate or arginine), this mechanism being a means of regulating the enzyme activity in vivo. In the direction of octopine oxidation, the octopus enzyme showed a much higher affinity for octopine (Km = 0.8 mM) than did the squid enzyme (Km = 4.4 mM), suggesting that it may be better geared for reconverting octopine to arginine and pyruvate after anaerobic bursts of muscle activity.

Biochemical changes in rat endometrium induced by oestrogens

1981 ◽  
Vol 96 (3) ◽  
pp. 382-388 ◽  
Author(s):  
Marta Jelínková ◽  
J. Jelínek ◽  
J. van der Vies
Keyword(s):  
Marked Effect ◽  
Specific Activity ◽  
Krebs Cycle ◽  
Regression Curve ◽  
Adult Rats ◽  
Oestrogen Treatment ◽  
Linear Dose ◽  
Cycle Plays ◽  
Wet Weight ◽  
Ovariectomized Adult Rats

Abstract. The effect of oestradiol (Oe2) or ethinyloestradiol (EOe) on several enzymes, soluble protein and DNA in the endometrium was studied in ovariectomized adult rats. The most marked effect of Oe2 was the increase in the activity of lactate dehydrogenase (LDH). If the treatment was started on the day of ovariectomy, it took approximately 8 days to reach a constant elevation. If the administration of Oe2 was started one week after castration, the maximum response was seen after 4 days of treatment. Using the latter schedule a linear dose-reponse regression curve of LDH/DNA was obtained with λ = 0.086. This parameter is considered suitable for the comparison of the effects of different oestrogens. Administration of Oe2 for 8 days beginning on the day of ovariectomy gave no linear dose-response curve of LDH/DNA. Administration of EOe caused a very marked increase of the specific activity of pyruvate kinase, LDH, M-type LDH and some slight, but significant decreases in isocitrate dehydrogenase, glutamate dehydrogenase, acid phosphatase and alkaline phosphatase. The changes of β-glucuronidase were only slight. The content of DNA per wet weight of endometrium decreased after oestrogen treatment, the protein content remained reasonably constant. It is concluded that, after stimulation with oestrogen, the rat endometrium produces the energy needed for its own growth mainly via anaerobic glycolysis and that the Krebs cycle plays a relatively small role.

Levels of Stable Zn and 65Zn in Crassostrea virginica from North Carolina

1970 ◽  
Vol 27 (1) ◽  
pp. 47-57 ◽  
Author(s):  
Douglas A. Wolfe
Keyword(s):  
North Carolina ◽  
Crassostrea Virginica ◽  
Half Life ◽  
Specific Activity ◽  
Adductor Muscle ◽  
Animal Tissues ◽  
Wet Weight ◽  
Zinc Levels

The concentration of zinc in oysters was highly variable — samples from relatively unpolluted estuaries of North Carolina contained, on the average, 85–245 ppm zinc, based on wet weight. Internal tissues, like adductor muscle and pericardial sac, had zinc levels less than half those of external tissues but zinc was nonetheless distributed uniformly throughout the animal tissues. During 1964–66, North Carolina oysters contained 2–20 pCi 65Zn from fallout per 100 g wet weight. Specific activity of 65Zn in these oysters during 1965–66 was in the range 90–300 pCi/g Zn, and was declining with an apparent half-life of 276 days.

scholarly journals Study of Extraction and Enzymatic Properties of Cell-Envelope Proteinases from a Novel Wild Lactobacillus plantarum LP69

Catalysts ◽  
2018 ◽  
Vol 8 (8) ◽  
pp. 325 ◽  
Author(s):  
He Chen ◽  
Jie Huang ◽  
Binyun Cao ◽  
Li Chen ◽  
Na Song ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Lactobacillus Plantarum ◽  
Industrial Development ◽  
Extraction Efficiency ◽  
Specific Activity ◽  
Cell Envelope ◽  
Serine Proteinase ◽  
Kinetic Studies ◽  
Predicted Values ◽  
Hydrolyze Whey Protein

Lactobacilli cell-envelope proteinases (CEPs) have been widely used in the development of new streams of blockbuster nutraceuticals because of numerous biopharmaceutical potentials; thus, the development of viable methods for CEP extraction and the improvement of extraction efficiency will promote their full-scale application. In this study, CEP from a novel wild Lactobacillus plantarum LP69 was released from cells by incubating in calcium-free buffer. The extraction conditions of CEP were optimized by response surface methodology with the enzyme activity and specific activity as the detective marker. The optimal extraction conditions were: time of 80 min, temperature of 39 °C and buffer pH of 6.5. Under these conditions, enzyme activity and specific activity were (23.94 ± 0.86) U/mL and (1.37 ± 0.03) U/mg, respectively, which were well matched with the predicted values (22.12 U/mL and 1.36 U/mg). Optimal activity of the crude CEP occurred at pH 8.0 and 40 °C. It is a metallopeptidase, activated by Ca2+, inhibited by Zn2+ and ethylene-diamine-tetra-acetic acid, and a serine proteinase which is inhibited by phenylmethylsulfonyl fluoride. Kinetic studies showed that CEP from LP69 could hydrolyze whey protein, lactoglobulin and casein. Our study improves the extraction efficiency of CEPs from LP69, providing the reference for their industrial development.

Large-scale production of citrate synthase from a cloned gene

1982 ◽  
Vol 60 (12) ◽  
pp. 1143-1147 ◽  
Author(s):  
Harry W. Duckworth ◽  
Alexander W. Bell
Keyword(s):  
Escherichia Coli ◽  
Large Scale ◽  
Citrate Synthase ◽  
Specific Activity ◽  
Tetracycline Resistance ◽  
Scale Production ◽  
Ampicillin Resistance ◽  
Colicin E1 ◽  
Large Scale Production ◽  
Cloned Gene

Starting with a colicin E1 resistance recombinant plasmid which contains gltA, the gene for citrate synthase in Escherichia coli, we have constructed an ampicillin-resistance plasmid containing the gltA region as a 2.9-kilobase-pair insert in the tetracycline-resistance region of pBR322. Escherichia coli HB101 harbouring this plasmid, when grown on rich medium containing ampicillin, contains citrate synthase as about 8% of its soluble protein. The enzyme has been purified from this rich source and is identical to the chromosomal enzyme prepared previously in every property tested, except for specific activity, which is 64 U∙mg−1 as compared with 45–50 U∙mg−1 previously obtained. The N-terminal sequences of both enzymes are reported, and they are identical up to residue 16 at least. The overall yield of pure enzyme, starting with the cells grown in 15 L of medium, is 600–800 mg.

Binding of 3-phosphoglycerate leads to both activation and stabilisation of ADP-glucose pyrophosphorylase from apple leaves

2005 ◽  
Vol 32 (9) ◽  
pp. 839
Author(s):  
Rui Zhou ◽  
Lailiang Cheng
Keyword(s):  
Heat Treatment ◽  
Thermal Stability ◽  
Enzyme Activity ◽  
Inorganic Phosphate ◽  
Thermal Inactivation ◽  
Specific Activity ◽  
Apple Leaves ◽  
Apparent Homogeneity ◽  
Adp Glucose Pyrophosphorylase ◽  
High Concentrations

Apple leaf ADP-glucose pyrophosphorylase was purified 1436-fold to apparent homogeneity with a specific activity of 58.9 units mg–1. The enzyme was activated by 3-phosphoglycerate (PGA) and inhibited by inorganic phosphate (Pi) in the ADPG synthesis direction. In the pyrophosphorolytic direction, however, high concentrations of PGA (> 2.5 mm) inhibited the enzyme activity. The enzyme was resistant to thermal inactivation with a T0.5 (temperature at which 50% of the enzyme activity is lost after 5 min incubation) of 52°C. Incubation with 2 mm PGA or 2 mm Pi increased T0.5 to 68°C. Incubation with 2 mm dithiothreitol (DTT) decreased T0.5 to 42°C, whereas inclusion of 2 mm PGA in the DTT incubation maintained T0.5 at 52°C. DTT-induced decrease in thermal stability was accompanied by monomerisation of the small subunits. Presence of PGA in the DTT incubation did not alter the monomerisation of the small subunits of the enzyme induced by DTT. These findings indicate that binding of PGA renders apple leaf AGPase with a conformation that is not only more efficient in catalysis but also more stable to heat treatment. The physiological significance of the protective effect of PGA on thermal inactivation of AGPase is discussed.

5α-Reductase activity in stroma and epithelium of rat prostate and epididymis. A contribution to elucidation of the mechanism for development of hyperplastic growth of prostatic tissue

1983 ◽  
Vol 103 (2) ◽  
pp. 273-281 ◽  
Author(s):  
Ole Djøseland ◽  
Nicholas Bruchovsky ◽  
Paul S. Rennie ◽  
Navdeep Otal ◽  
Sian Høglo
Keyword(s):  
Enzyme Activity ◽  
Specific Activity ◽  
Reductase Activity ◽  
Major Change ◽  
Growth Stimulation ◽  
Total Activity ◽  
Prostatic Tissue ◽  
Ventral Prostate ◽  
Total Enzyme Activity ◽  
Abnormal Growth

Abstract. The 5α-reductase activity was assayed in homogenates of stroma and epithelium in the rat ventral prostate and epididymis. Samples consisting of 0.3 mg/ml tissue protein in TES buffer, pH 7.0 were incubated at 37°C for 30 min in the presence of 50 nm [1,2-3H]testosterone and a NADPH-generating system started with 5 × 10−4 m NADP. The yield of 5α-reduced metabolites, as established by using thin-layer chromatography, gave an estimate of enzyme activity. Whereas the specific activity of 5α-reductase was highest in prostatic stroma and epididymal epithelium, most of the total enzyme activity was associated with the epithelium in both the prostate and epididymis. The effect of dihydrotestosterone on specific activity of 5α-reductase was studied by administering the hormone to 7-day castrated rats. In prostate, the specific activity of both stromal and epithelium forms of the enzyme reached a maximum after 4 days of treatment. In epididymis only the epithelial form of 5α-reductase underwent a major change in specific activity, the latter peaking after 8–12 days of treatment. Furthermore, while the total activity of 5α-reductase in the prostatic tissue fractions could be induced by as much as 4-fold the normal control values, the epididymal enzyme could not be induced above the normal level either in the stroma or the epithelium. This may explain the relative resistance of epididymis to abnormal growth stimulation under the influence of hormones.

scholarly journals Rat uterine microbiochemistry: metabolic enzyme activities stimulated by 17-beta-estradiol are localized in epithelial cells.

1987 ◽  
Vol 35 (6) ◽  
pp. 657-662 ◽  
Author(s):  
J P Holt ◽  
E Rhe
Keyword(s):  
Enzyme Activity ◽  
Smooth Muscle ◽  
Epithelial Cells ◽  
Dose Response ◽  
Enzyme Activities ◽  
Time Course ◽  
Citrate Synthase ◽  
Ovariectomized Rats ◽  
Similar Response ◽  
Estradiol Treatment

Lactate dehydrogenase (LDH; EC 1.1.1.27), citrate synthase (CS; EC 4.1.3.7), and beta-hydroxyacyl-CoA-dehydrogenase (beta-OH-acyl-CoA-DH; EC 1.1.1.35) activities were determined in each of the three major cell types of rat uterus, i.e., epithelial, stromal, and smooth muscle, using quantitative microanalytical techniques. Adult ovariectomized rats were treated with 17-beta-estradiol to determine the time course and dose response (0.025-50 micrograms/300-g rat) effect of estrogen on enzyme activity of each type of uterine cell. The use of "oil well" and enzyme-cycling microtechniques to determine the time course and the dose responses of enzyme activity changes required microassays involving 1595 microdissected single cell specimens. Estradiol treatment increased epithelial LDH, CS and beta-OH-acyl-CoA-DH activity but had no effect on these enzymes in the stroma or in smooth muscle cells. The estradiol-stimulated peak enzyme activities on Day 4 in the intervention group are compared with those in the ovariectomized rat controls as follows: LDH, 44.5 +/- 3.5 vs 22.3 +/- 3.9; CS, 3.5 +/- 0.2 vs 1.5 +/- 0.6; beta-OH-acyl-CoA-H, 3.5 +/- 0.32 vs 2.2 +/- 0.2 (mean +/- standard deviation; mol/kg/hr). Stromal cell activities (LDH, 7.4 +/- 1.0; CS, 1.2 +/- 0.2; beta-OH-acyl-CoA-DH, 0.9 +/- 0.1) were significantly lower than epithelial cell levels and were similar to smooth muscle levels. Therefore, even in the ovariectomized animal epithelial cells have markedly higher metabolic activity compared with adjacent cells. The enzyme activities are expressed as moles of substrate reacting per kilogram of dry weight per hour. All three enzymes exhibited a 17-beta-estradiol-induced dose response between 0.025-0.15 micrograms/300-g rat. The three enzymes studied all had similar response patterns to estrogen. The effect of estradiol was restricted to epithelial cells, with enzyme activities increasing to maximal levels after approximately 96 hr of hormone treatment. This study therefore not only confirms the specific and differential metabolic responses of uterine cells to estradiol treatment, but clearly demonstrates that marked metabolic differences exist between epithelial cells and stromal or smooth muscle uterine cells.

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