Biosynthesis of polyglutamates of folates

1986 ◽  
Vol 64 (7) ◽  
pp. 667-674 ◽  
Author(s):  
K. G. Scrimgeour
Keyword(s):  
Current Knowledge ◽  
Competitive Inhibitor ◽  
High Dose ◽  
Beef Liver ◽  
Folylpolyglutamate Synthetase ◽  
Low Concentrations ◽  
Alternate Substrate ◽  
Pteroic Acid ◽  
Rescue Agent ◽  
Addition Pathway

Folylpolyglutamate synthetase (FPGS) catalyzes the synthesis of the poly-γ-glutamate forms of tetrahydrofolate and its coenzyme adducts, as well as of the folate-analogue drugs. This paper reviews current knowledge of the preparations of FPGS from mammalian sources (rat, hog, mouse, and beef liver). Kinetic constants for the substrates and activators of FPGS are compared. Tetrahydrofolate and 5-formyltetrahydrofolate are excellent substrates for the enzyme. The Km values for the antifolates and their 7-hydroxy metabolites are much higher than those for the tetrahydrofolates. Aminopterin has higher activity with FPGS than does methotrexate, which partially explains its greater toxicity. 5-Formyltetrahydrofolate, which is used as a rescue agent in high-dose methotrexate-rescue chemotherapy, is a better alternate substrate of FPGS than is methotrexate and therefore is a potent competitive inhibitor of the glutamylation of methotrexate. Thus, low concentrations of the rescue agent prevent formation of cytotoxic polyglutamates of methotrexate. The pathway of the reaction is the addition of a glutamate residue to the terminal γ-carboxyl of the pteridine substrate. That longer folylpolyglutamates are poorer substrates possibly is a result of this addition pathway. Pteroic acid activates FPGS by lowering the Km value of the pteridine substrate. It also greatly increases the activity of the synthetase at physiological pH values.

Activation of folylpolyglutamate synthetase by pteroic acid

1985 ◽  
Vol 63 (7) ◽  
pp. 777-779 ◽  
Author(s):  
P. J. Vickers ◽  
R. Di Cecco ◽  
Z. B. Pristupa ◽  
K. G. Scrimgeour
Keyword(s):  
Conformational Change ◽  
Ph Optimum ◽  
Beef Liver ◽  
Folylpolyglutamate Synthetase ◽  
Ph Values ◽  
Neutral Ph ◽  
Pteroic Acid ◽  
Stimulation Of

The glutamylation of methotrexate catalyzed by beef liver folylpolyglutamate synthetase (FPGS) is activated by addition of pteroic acid. Pteroic acid causes greater stimulation of FPGS, including glutamylation of tetrahydrofolate, at neutral pH values (i.e., below the pH optimum of 8.4). We have attributed this activation to a conformational change of FPGS induced by pteroic acid.

Serum Cholinesterase Variants: Examination of Several Differential Inhibitors, Salts, and Buffers Used to Measure Enzyme Activity

1971 ◽  
Vol 17 (3) ◽  
pp. 183-191 ◽  
Author(s):  
Philip J Garry
Keyword(s):  
Enzyme Activity ◽  
Sodium Fluoride ◽  
Phosphate Buffer ◽  
Divalent Cations ◽  
Kinetic Studies ◽  
Competitive Inhibitor ◽  
Serum Cholinesterase ◽  
Low Concentrations

Abstract Dibucaine, used as a differential inhibitor with acetyl-, propionyl-, and butyrylthiocholine as substrate, clearly identified the "usual" and "atypical" serum cholinesterases. Succinylcholine was also used successfully as a differential inhibitor with butyrylthiocholine as substrate. Sodium fluoride, used as a differential inhibitor, gave conflicting results, depending on whether Tris or phosphate buffer was used in the assay. Mono- and divalent cations (NaCl, KCl, MgCl2, CaCl2, and BaCl2) activated the "usual" and inhibited the "atypical" enzyme at low concentrations. The "usual" enzyme had the same activity in 0.05 mol of Tris or phosphate buffer per liter, while the heterozygous and "atypical" enzymes showed 12 and 42% inhibition, respectively, when assayed in the phosphate buffer. Kinetic studies showed the phosphate acted as a competitive inhibitor of "atypical" enzyme. Km values, determined for "usual" and "atypical" enzymes, were 0.057 and 0.226 mmol/liter, respectively, with butyrylthiocholine as substrate.

scholarly journals Mitochondrial adenosine triphosphatase from human placenta--inhibition by free magnesium ions of ITP hydrolysis.

1994 ◽  
Vol 41 (1) ◽  
pp. 39-44 ◽  
Author(s):  
Z Aleksandrowicz
Keyword(s):  
Competitive Inhibition ◽  
Competitive Inhibitor ◽  
Hill Coefficient ◽  
Negative Cooperativity ◽  
Magnesium Ions ◽  
Beef Liver ◽  
Bicarbonate Ions ◽  
The Hill ◽  
Kinetics Of ◽  
Free Magnesium

The effects of Mg2+ and bicarbonate on the kinetics of ITP hydrolysis by soluble ATPase (F1) from human placental mitochondria were studied. Increasing amounts of Mg2+ at fixed ITP concentration, caused a marked activation of F1 followed by inhibition at higher Mg2+ concentration. The appropriate substrate for the mitochondrial F1 seems to be the MgITP complex as almost no ITP was hydrolysed in the absence of magnesium. Mg2+ behaved as a competitive inhibitor towards the MgITP complex. In this respect the human placental enzyme differ from that from other sources such as yeast, beef liver or rat liver. The linearity of the plot presenting competitive inhibition by free Mg2+ of MgITP hydrolysis (in the presence of activating bicarbonate anion) suggests that both Mg2+ and MgITP bind to the same catalytic site (Km(MgITP) = 0.46 mM, Ki(Mg) = 4 mM). When bicarbonate was absent in the ITPase assay, placental F1 exhibited apparent negative cooperativity in the presence of 5 mM Mg2+, just as it did with MgATP as a substrate under similar conditions. Bicarbonate ions eliminated the negative cooperativity with respect to ITP (as the Hill coefficient of 0.46 was brought to approx. 1), and thus limited inhibition by free Mg2+. The results presented suggest that the concentration of free magnesium ions may be an important regulatory factor of the human placental F1 activity.

scholarly journals Back to the Future: Very High-Energy Electrons (VHEEs) and Their Potential Application in Radiation Therapy

Cancers ◽  
2021 ◽  
Vol 13 (19) ◽  
pp. 4942
Author(s):  
Maria Grazia Ronga ◽  
Marco Cavallone ◽  
Annalisa Patriarca ◽  
Amelia Maia Leite ◽  
Pierre Loap ◽  
...  
Keyword(s):  
Radiation Therapy ◽  
Current Knowledge ◽  
High Energy ◽  
High Dose Rate ◽  
Point Of View ◽  
High Dose ◽  
Dose Rates ◽  
High Energy Electrons ◽  
Very High Energy ◽  
Very High

The development of innovative approaches that would reduce the sensitivity of healthy tissues to irradiation while maintaining the efficacy of the treatment on the tumor is of crucial importance for the progress of the efficacy of radiotherapy. Recent methodological developments and innovations, such as scanned beams, ultra-high dose rates, and very high-energy electrons, which may be simultaneously available on new accelerators, would allow for possible radiobiological advantages of very short pulses of ultra-high dose rate (FLASH) therapy for radiation therapy to be considered. In particular, very high-energy electron (VHEE) radiotherapy, in the energy range of 100 to 250 MeV, first proposed in the 2000s, would be particularly interesting both from a ballistic and biological point of view for the establishment of this new type of irradiation technique. In this review, we examine and summarize the current knowledge on VHEE radiotherapy and provide a synthesis of the studies that have been published on various experimental and simulation works. We will also consider the potential for VHEE therapy to be translated into clinical contexts.

scholarly journals Alcohol Hangover Slightly Impairs Response Selection but not Response Inhibition

2019 ◽  
Vol 8 (9) ◽  
pp. 1317 ◽  
Author(s):  
Antje Opitz ◽  
Jan Hubert ◽  
Christian Beste ◽  
Ann-Kathrin Stock
Keyword(s):  
Cognitive Control ◽  
Response Inhibition ◽  
Response Selection ◽  
Current Knowledge ◽  
Alcohol Intoxication ◽  
Heavy Drinking ◽  
High Dose ◽  
Automatic Response ◽  
Alcoholic Drinks ◽  
Alcohol Hangover

Alcohol hangover commonly occurs after an episode of heavy drinking. It has previously been demonstrated that acute high-dose alcohol intoxication reduces cognitive control, while automatic processes remain comparatively unaffected. However, it has remained unclear whether alcohol hangover, as a consequence of binge drinking, modulates the interplay between cognitive control and automaticity in a comparable way. Therefore, the purpose of this study was to investigate the effects of alcohol hangover on controlled versus automatic response selection and inhibition. N = 34 healthy young men completed a Simon Nogo task, once sober and once hungover. Hangover symptoms were experimentally induced by a standardized administration of alcoholic drinks (with high congener content) on the night before the hangover appointment. We found no significant hangover effects, which suggests that alcohol hangover did not produce the same functional deficits as an acute high-dose intoxication. Yet still, add-on Bayesian analyses revealed that hangover slightly impaired response selection, but not response inhibition. This pattern of effects cannot be explained with the current knowledge on how ethanol and its metabolite acetaldehyde may modulate response selection and inhibition via the dopaminergic or GABAergic system.

scholarly journals Roles of IFN-γ in tumor progression and regression: a review

2020 ◽  
Vol 8 (1) ◽  
Author(s):  
Dragica Jorgovanovic ◽  
Mengjia Song ◽  
Liping Wang ◽  
Yi Zhang
Keyword(s):  
Immune Response ◽  
Tumor Progression ◽  
Tumor Regression ◽  
Current Knowledge ◽  
Checkpoint Inhibitors ◽  
Interferon Γ ◽  
High Dose ◽  
Main Body ◽  
Significant Research ◽  
Ifn Γ

Abstract Background Interferon-γ (IFN-γ) plays a key role in activation of cellular immunity and subsequently, stimulation of antitumor immune-response. Based on its cytostatic, pro-apoptotic and antiproliferative functions, IFN-γ is considered potentially useful for adjuvant immunotherapy for different types of cancer. Moreover, it IFN-γ may inhibit angiogenesis in tumor tissue, induce regulatory T-cell apoptosis, and/or stimulate the activity of M1 proinflammatory macrophages to overcome tumor progression. However, the current understanding of the roles of IFN-γ in the tumor microenvironment (TME) may be misleading in terms of its clinical application. Main body Some researchers believe it has anti-tumorigenic properties, while others suggest that it contributes to tumor growth and progression. In our recent work, we have shown that concentration of IFN-γ in the TME determines its function. Further, it was reported that tumors treated with low-dose IFN-γ acquired metastatic properties while those infused with high dose led to tumor regression. Pro-tumorigenic role may be described through IFN-γ signaling insensitivity, downregulation of major histocompatibility complexes, upregulation of indoleamine 2,3-dioxygenase, and checkpoint inhibitors such as programmed cell death ligand 1. Conclusion Significant research efforts are required to decipher IFN-γ-dependent pro- and anti-tumorigenic effects. This review discusses the current knowledge concerning the roles of IFN-γ in the TME as a part of the complex immune response to cancer and highlights the importance of identifying IFN-γ responsive patients to improve their sensitivity to immuno-therapies.

scholarly journals The enzymology of short-chain fatty acyl-coenzyme A synthetase from seeds of Pinus radiata. Kinetic studies and a proposed reaction mechanism

1974 ◽  
Vol 137 (3) ◽  
pp. 435-442 ◽  
Author(s):  
Owen A. Young ◽  
John W. Anderson
Keyword(s):  
Fatty Acid ◽  
Pinus Radiata ◽  
Kinetic Studies ◽  
Single Species ◽  
Competitive Inhibitor ◽  
Fatty Acyl ◽  
Short Chain ◽  
Low Concentrations ◽  
High Concentrations ◽  
Acyl Coenzyme A

1. Short-chain fatty acyl-CoA synthetase from seeds of Pinus radiata was examined by acetate- and propionate-dependent PPi–ATP exchange. Reaction mixtures came to equilibrium almost instantly as judged by rates of exchange and analysis of an incubation mixture. 2. The activity of the enzyme was correlated with the concentration of MgP2O72- but not with the concentration of Mg2+, as judged by PPi–ATP exchange and fatty acyl AMP-dependent synthesis of ATP in the presence of PPi. In PPi–ATP exchange assays, no clear relationship between activity and any single species of ATP was apparent. 3. High concentrations of fatty acid inhibited PPi–ATP exchange. PPi–dATP exchange was less than PPi–ATP exchange at low concentrations of fatty acid, but at higher concentrations PPi–dATP exchange exceeded PPi–ATP exchange. The rate of synthesis of fatty acyl-CoA in the presence of dATP was less than with ATP. 4. ATP and propionate inhibited the synthesis of ATP from propionyl-AMP and PPi. The inhibition by ATP was competitive with respect to propionyl-AMP and non-competitive with respect to PPi. The inhibition by propionate was non-competitive with respect to propionyl-AMP and PPi. 5. AMP was a competitive inhibitor of propionyl-AMP-dependent synthesis of ATP and competitively inhibited propionate-dependent PPi–ATP exchange when ATP was the variable substrate. 6. It was concluded that the first partial reaction catalysed by the enzyme is ordered; ATP is the first substrate to react with the enzyme and PPi is probably the only product released.

scholarly journals Chick osteoblasts contain fluoride-sensitive acid phosphatase activity.

1988 ◽  
Vol 36 (9) ◽  
pp. 1175-1180 ◽  
Author(s):  
M W Lundy ◽  
K H Lau ◽  
H C Blair ◽  
D J Baylink
Keyword(s):  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Acid Phosphatase Activity ◽  
Bone Matrix ◽  
Competitive Inhibitor ◽  
Cellular Origin ◽  
Embryonic Chicken ◽  
Low Concentrations

We used histological and biochemical methods to determine the cellular origin of bone matrix fluoride-sensitive acid phosphatase in chicken bone. Embryonic chicken calvariae were embedded in plastic and sections stained for acid phosphatase at various concentrations of substrate and fluoride. Acid phosphatase activity was observed in osteoblasts and osteoclasts but not in fibroblasts. Striking inhibition of osteoblastic acid phosphatase occurred at 100 microM fluoride, a concentration that had no apparent effect on osteoclastic acid phosphatase. Inhibition of osteoblastic and osteoclastic acid phosphatase by fluoride was also examined using extracts of embryonic chicken calvarial cells, mouse osteoblasts (MC3T3-El cell line), and purified chick osteoclasts, respectively. Fluoride is a partial competitive inhibitor of both chicken and mouse osteoblastic acid phosphatases, with apparent inhibition constants of 10-100 microM. These concentrations of fluoride correspond to those that increase bone formation in vitro and in vivo. In contrast, the apparent inhibition constant for fluoride of osteoclastic acid phosphatase was much higher (i.e., 0.5 mM). In summary, this study demonstrates that chicken osteoblasts contain an acid phosphatase that is sensitive to inhibition by low concentrations (i.e., microM) of fluoride.

scholarly journals Inhibition of Hepatitis C Virus Replication by GS-6620, a PotentC-Nucleoside Monophosphate Prodrug

2014 ◽  
Vol 58 (4) ◽  
pp. 1930-1942 ◽  
Author(s):  
Joy Y. Feng ◽  
Guofeng Cheng ◽  
Jason Perry ◽  
Ona Barauskas ◽  
Yili Xu ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Nonstructural Protein ◽  
Competitive Inhibitor ◽  
High Dose ◽  
Diarrhea Virus ◽  
A Chain ◽  
Direct Acting ◽  
Hcv Ns5b

ABSTRACTAs a class, nucleotide inhibitors (NIs) of the hepatitis C virus (HCV) nonstructural protein 5B (NS5B) RNA-dependent RNA polymerase offer advantages over other direct-acting antivirals, including properties, such as pangenotype activity, a high barrier to resistance, and reduced potential for drug-drug interactions. We studied thein vitropharmacology of a novelC-nucleoside adenosine analog monophosphate prodrug, GS-6620. It was found to be a potent and selective HCV inhibitor against HCV replicons of genotypes 1 to 6 and against an infectious genotype 2a virus (50% effective concentration [EC50], 0.048 to 0.68 μM). GS-6620 showed limited activities against other viruses, maintaining only some of its activity against the closely related bovine viral diarrhea virus (EC50, 1.5 μM). The active 5′-triphosphate metabolite of GS-6620 is a chain terminator of viral RNA synthesis and a competitive inhibitor of NS5B-catalyzed ATP incorporation, withKi/Kmvalues of 0.23 and 0.18 for HCV NS5B genotypes 1b and 2a, respectively. With its unique dual substitutions of 1′-CN and 2′-C-Me on the ribose ring, the active triphosphate metabolite was found to have enhanced selectivity for the HCV NS5B polymerase over host RNA polymerases. GS-6620 demonstrated a high barrier to resistancein vitro. Prolonged passaging resulted in the selection of the S282T mutation in NS5B that was found to be resistant in both cellular and enzymatic assays (>30-fold). Consistent with itsin vitroprofile, GS-6620 exhibited the potential for potent anti-HCV activity in a proof-of-concept clinical trial, but its utility was limited by the requirement of high dose levels and pharmacokinetic and pharmacodynamic variability.

scholarly journals A synthesis of atmospheric mercury depletion event chemistry in the atmosphere and snow

2008 ◽  
Vol 8 (6) ◽  
pp. 1445-1482 ◽  
Author(s):  
A. Steffen ◽  
T. Douglas ◽  
M. Amyot ◽  
P. Ariya ◽  
K. Aspmo ◽  
...  
Keyword(s):  
Sea Ice ◽  
Current Knowledge ◽  
Kinetic Studies ◽  
Elemental Mercury ◽  
Atmospheric Mercury ◽  
The Arctic ◽  
Polar Regions ◽  
Environmental Media ◽  
Gaseous Elemental Mercury ◽  
Low Concentrations

Abstract. It was discovered in 1995 that, during the spring time, unexpectedly low concentrations of gaseous elemental mercury (GEM) occurred in the Arctic air. This was surprising for a pollutant known to have a long residence time in the atmosphere; however conditions appeared to exist in the Arctic that promoted this depletion of mercury (Hg). This phenomenon is termed atmospheric mercury depletion events (AMDEs) and its discovery has revolutionized our understanding of the cycling of Hg in Polar Regions while stimulating a significant amount of research to understand its impact to this fragile ecosystem. Shortly after the discovery was made in Canada, AMDEs were confirmed to occur throughout the Arctic, sub-Artic and Antarctic coasts. It is now known that, through a series of photochemically initiated reactions involving halogens, GEM is converted to a more reactive species and is subsequently associated to particles in the air and/or deposited to the polar environment. AMDEs are a means by which Hg is transferred from the atmosphere to the environment that was previously unknown. In this article we review Hg research taken place in Polar Regions pertaining to AMDEs, the methods used to collect Hg in different environmental media, research results of the current understanding of AMDEs from field, laboratory and modeling work, how Hg cycles around the environment after AMDEs, gaps in our current knowledge and the future impacts that AMDEs may have on polar environments. The research presented has shown that while considerable improvements in methodology to measure Hg have been made but the main limitation remains knowing the speciation of Hg in the various media. The processes that drive AMDEs and how they occur are discussed. As well, the role that the snow pack and the sea ice play in the cycling of Hg is presented. It has been found that deposition of Hg from AMDEs occurs at marine coasts and not far inland and that a fraction of the deposited Hg does not remain in the same form in the snow. Kinetic studies undertaken have demonstrated that bromine is the major oxidant depleting Hg in the atmosphere. Modeling results demonstrate that there is a significant deposition of Hg to Polar Regions as a result of AMDEs. Models have also shown that Hg is readily transported to the Arctic from source regions, at times during springtime when this environment is actively transforming Hg from the atmosphere to the snow and ice surfaces. The presence of significant amounts of methyl Hg in snow in the Arctic surrounding AMDEs is important because this species is the link between the environment and impacts to wildlife and humans. Further, much work on methylation and demethylation processes has occurred but these processes are not yet fully understood. Recent changes in the climate and sea ice cover in Polar Regions are likely to have strong effects on the cycling of Hg in this environment; however more research is needed to understand Hg processes in order to formulate meaningful predictions of these changes.

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