scholarly journals Properties of freshly purified and thiol-treated spinach chloroplast fructose bisphosphatase

1980 ◽  
Vol 185 (3) ◽  
pp. 689-693 ◽  
Author(s):  
S A Charles ◽  
B Halliwell
Keyword(s):  
Inorganic Phosphate ◽  
Physiological Significance ◽  
Fructose Bisphosphatase ◽  
Spinach Chloroplast ◽  
Significant Rate ◽  
Chloroplast Stroma ◽  
Fructose 1,6 Bisphosphate

Freshly purified spinach chloroplast fructose bisphosphatase is powerfully inhibited by inorganic phosphate competitively with respect to its substrate fructose 1,6-bisphosphate. The concentrations of phosphate and substrate in the chloroplast stroma are such that the enzyme in this form could not operate at a significant rate in vivo. Incubation of the enzyme with dithiothreitol for 24 h decreases the Km for fructose 1,6-bisphosphate from 0.8 to 0.033 mM, decreases the Km for Mg2+ from 9 to 2 mM and substantially alleviates inhibition by inorganic phosphate. The physiological significance of thiol activation of the enzyme is discussed.

Regulation of Fructose 1,6-Bisphosphatase Activity of Chlorella by Mole Mass Change

1996 ◽  
Vol 51 (9-10) ◽  
pp. 639-645 ◽  
Author(s):  
N. Grotjohann
Keyword(s):  
Enzyme Regulation ◽  
Fold Increase ◽  
Substrate Affinity ◽  
Kinetic Properties ◽  
Enzyme Form ◽  
Phosphorylated Compounds ◽  
Fructose 1,6 Bisphosphatase ◽  
Chloroplast Stroma ◽  
Fructose 1,6 Bisphosphate

Fast protein liquid chromatography on Superose 6 of partially purified FBPase II from Chlorella reveals a 1350 kDa-form at pH 6.0 and a 67 kDa-form at pH 8.5. Treatment of the large enzyme form with 5mᴍ concentrations of Mg2+, F1,6P2, DTT or ATP leads to dissociation into smaller ones of 215 -470 kDa. Aggregation/dissoziation is a reversible process, as has been shown for the effect of F1,6P2 and of pH, by rechromatography. The change in mole mass results in alterations of the activitiy and of the kinetic properties of the enzyme forms, obtained. Dissociation results in a 4 - 6 fold increase in activity, as can be shown for F1,6P2-treated samples. Halfsaturation constants, as well as the degree of cooperativity of the 67- and the 1350- kDa form, are different for substrate affinity, activation by Mg2+ and DTT, and for inhibition by ATP. Both enzyme forms hydrolyse fructose 1,6 bisphosphate and seduheptulose 1,7 bisphosphate better than other phosphorylated compounds. The ratio of F1,6P2- to SDP-cleavage is 100:58 for the small enzyme form and 100: 84 for the large one. Activation of FBPase II in the light and inactivation in the dark is discussed on the basis of different oligomeric forms of the enzyme, generated by changes in the concentration of intermediates and effectors in the chloroplast stroma, leading to dissociation or aggregation. The conclusion is drawn that oligomerization of key enzymes, resulting in enzyme forms with different activities and different kinetic properties, might provide an effective mechanism for enzyme regulation in vivo

Studies with GIP/Ins cells indicate secretion by gut K cells is KATPchannel independent

2003 ◽  
Vol 284 (5) ◽  
pp. E988-E1000 ◽  
Author(s):  
Song Yan Wang ◽  
Maggie M.-Y. Chi ◽  
Lin Li ◽  
Kelle H. Moley ◽  
Burton M. Wice
Keyword(s):  
Tricarboxylic Acid Cycle ◽  
Calcium Mobilization ◽  
Enteroendocrine Cells ◽  
Glycolytic Flux ◽  
K Cells ◽  
Antibody Staining ◽  
Methyl Pyruvate ◽  
Fructose 1,6 Bisphosphate ◽  
Insulinoma Cells

K cells are a subpopulation of enteroendocrine cells that secrete glucose-dependent insulinotropic polypeptide (GIP), a hormone that promotes glucose homeostasis and obesity. Therefore, it is important to understand how GIP secretion is regulated. GIP-producing (GIP/Ins) cell lines secreted hormones in response to many GIP secretagogues except glucose. In contrast, glyceraldehyde and methyl pyruvate stimulated hormone release. Measurements of intracellular glucose 6-phosphate, fructose 1,6-bisphosphate, and pyruvate levels, as well as glycolytic flux, in glucose-stimulated GIP/Ins cells indicated that glycolysis was not impaired. Analogous results were obtained using glucose-responsive MIN6 insulinoma cells. Citrate levels increased similarly in glucose-treated MIN6 and GIP/Ins cells. Thus pyruvate entered the tricarboxylic acid cycle. Glucose and methyl pyruvate stimulated 1.4- and 1.6-fold increases, respectively, in the ATP-to-ADP ratio in GIP/Ins cells. Glyceraldehyde profoundly reduced, rather than increased, ATP/ADP. Thus nutrient-regulated secretion is independent of the ATP-dependent potassium (KATP) channel. Antibody staining of mouse intestine demonstrated that enteroendocrine cells producing GIP, glucagon-like peptide-1, CCK, or somatostatin do not express detectable levels of inwardly rectifying potassium (Kir) 6.1 or Kir 6.2, indicating that release of these hormones in vivo may also be KATPchannel independent. Conversely, nearly all cells expressing chromogranin A or substance P and ∼50% of the cells expressing secretin or serotonin exhibited Kir 6.2 staining. Compounds that activate calcium mobilization were potent secretagogues for GIP/Ins cells. Secretion was only partially inhibited by verapamil, suggesting that calcium mobilization from intracellular and extracellular sources, independent from KATPchannels, regulates secretion from some, but not all, subpopulations of enteroendocrine cells.

Control of glycolysis in vertebrate skeletal muscle during exercise

1996 ◽  
Vol 270 (4) ◽  
pp. R821-R829 ◽  
Author(s):  
U. Krause ◽  
G. Wegener
Keyword(s):  
Gastrocnemius Muscle ◽  
Rana Temporaria ◽  
High Capacity ◽  
Repeated Exercise ◽  
Frog Rana Temporaria ◽  
Time Courses ◽  
Fructose 1,6 Bisphosphate ◽  
Assay Conditions

The gastrocnemius muscle of the frog (Rana temporaria) has a high capacity for anaerobic glycolysis from glycogen. Glycolytic metabolites and effectors of phosphofructokinase, particularly the hexose bisphosphates, were followed in muscle during exercise (swimming between 5 s and 5 min), recovery (rest for up to 2 h after 5 min of swimming), and repeated exercise (swimming for up to 60 s after 2 h of recovery). Glycogen phosphorylase and phosphofructokinase were swiftly activated with exercise. The hexose bisphosphates followed markedly different time courses. Fructose 1,6-bisphosphate was transiently increased in both exercise and repeated exercise. This appears to be an effect rather than a cause of phosphofructokinase activation. Glucose 1,6-biphosphate was accumulated only while phosphofructokinase was active and was unchanged at other times. Fructose 2,6-biphosphate showed a 10-fold transient increase on exercise in rested frogs, almost disappeared from the muscle during recovery, and did not change during repeated exercise. Fructose 2,6-biphosphate is a potent activator of phosphofructokinase in vitro under near physiological assay conditions, and it may serve this function also in vivo during exercise. Glucose 1,6-biphosphate could be an activator of phosphofructokinase in repeated exercise when fructose 2,6-biphosphate is not available.

Effects of some anti-inflammatory drugs on 12 blood constituents: protocol for the study of in vivo effects of drugs.

1985 ◽  
Vol 31 (7) ◽  
pp. 1141-1143 ◽  
Author(s):  
Z Jelić-Ivanović ◽  
S Spasić ◽  
N Majkić-Singh ◽  
P Todorović
Keyword(s):  
Uric Acid ◽  
Acetylsalicylic Acid ◽  
Total Protein ◽  
Inorganic Phosphate ◽  
Drug Effects ◽  
Urea Nitrogen ◽  
Blood Constituents ◽  
Inflammatory Drugs ◽  
In Vivo Effects

Abstract We investigated the in vivo effects of acetylsalicylic acid, diclofenac, indomethacin, ibuprofen, and ketoprofen on the concentrations of various blood constituents. Total protein, glucose, calcium, and inorganic phosphate were not significantly affected by any of these drugs. Ketoprofen had no definite influence on any constituent. Acetylsalicylic acid induced an increase in cholesterol, triglyceride, and iron; albumin, uric acid, and creatinine decreased with ibuprofen therapy. Urea nitrogen increased in patients treated with diclofenac or indomethacin. Our protocol for the study of in vivo drug effects is discussed.

scholarly journals Stimulation by Light of Rapid pH Regulation in the Chloroplast Stroma in Vivo as Indicated by CO2 Solubilization in Leaves

1995 ◽  
Vol 108 (3) ◽  
pp. 1059-1066 ◽  
Author(s):  
M. Hauser ◽  
H. Eichelmann ◽  
V. Oja ◽  
U. Heber ◽  
A. Laisk
Keyword(s):  
Ph Regulation ◽  
Chloroplast Stroma

31P-NMR studies of cerebral metabolic changes during graded hypoxia in newborn lambs

1987 ◽  
Vol 62 (4) ◽  
pp. 1569-1574 ◽  
Author(s):  
D. P. Younkin ◽  
L. C. Wagerle ◽  
B. Chance ◽  
J. Maria ◽  
M. Delivoria-Papadopoulos
Keyword(s):  
Steady State ◽  
Inorganic Phosphate ◽  
31P Nmr ◽  
Base Line ◽  
Nmr Studies ◽  
Steady State Kinetics ◽  
O2 Partial Pressure ◽  
Large Fluctuations ◽  
Early Phases

We measured cerebral phosphocreatine (PCr), inorganic phosphate (Pi), ATP, and intracellular pH (pHi) with in vivo phosphorus nuclear magnetic resonance (NMR) during 10- to 15-min periods of reversible hypoxic hypoxia in 20 newborn lambs (1–11 days). There was a significant correlation between arterial O2 partial pressure (PaO2) and the PCr/Pi ratio or pHi; however, between PaO2 130–33 mmHg, metabolite changes were not significant. PCr/Pi and pHi decreased significantly when PaO2 was lowered below 33 and 28 mmHg, respectively. After recovery, metabolite ratios and pHi returned to base-line values within 5 min. During the early phases of hypoxia and recovery, there were large fluctuations in metabolites and pHi, indicating that mitochondrial reactions were not in a steady state. After several minutes of hypoxia or recovery, PCr/Pi and pHi stabilized, suggesting steady state kinetics for mitochondrial respiration. NMR is extremely sensitive to changes in mitochondrial oxygenation, and stable PCr/Pi and pHi indicate that O2 tension in cerebral mitochondria of the newborn lamb is constant between PaO2 of 30 and 140 mmHg.

scholarly journals THE USE OF BISMUTH AS AN ELECTRON STAIN FOR NUCLEIC ACIDS

1963 ◽  
Vol 17 (1) ◽  
pp. 93-103 ◽  
Author(s):  
Peter Albersheim ◽  
Ursula Killias
Keyword(s):  
Nucleic Acids ◽  
Inorganic Phosphate ◽  
In Vitro Experiments ◽  
Root Tips ◽  
Onion Root ◽  
Cell Structures ◽  
Dividing Cells ◽  
Chromatin Material

Evidence is presented to show that bismuth combines in vitro with the phosphate of nucleic acids in a manner similar to its reaction with inorganic phosphate. When tested under similar conditions, protein exhibited no attraction for bismuth. The results of the in vitro experiments, which are of interest within themselves, may be indirectly applicable to in vivo staining. Dividing cells of onion root tips were fixed in OsO4, stained with bismuth, and examined in the electron microscope. The electron opacity of cell structures known to contain nucleic acids was enhanced by bismuth, while organelles known to lack appreciable quantities of DNA or RNA showed little, if any, change. Bismuth is particularly effective as a stain for the chromatin material during interphase and for the chromosomes during division.

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