scholarly journals Hydroxylamine Analogue of Agmatine: Magic Bullet for Arginine Decarboxylase

Biomolecules ◽  
2020 ◽  
Vol 10 (3) ◽  
pp. 406 ◽  
Author(s):  
Mervi T. Hyvönen ◽  
Tuomo A. Keinänen ◽  
Gulgina K. Nuraeva ◽  
Dmitry V. Yanvarev ◽  
Maxim Khomutov ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Cellular Proliferation ◽  
Arginine Decarboxylase ◽  
Magic Bullet ◽  
High Concentration ◽  
Polyamine Biosynthesis ◽  
E Coli ◽  
Growth Inhibitory ◽  
Inhibitory Analysis ◽  
Important Addition

The biogenic polyamines, spermine, spermidine (Spd) and putrescine (Put) are present at micro-millimolar concentrations in eukaryotic and prokaryotic cells (many prokaryotes have no spermine), participating in the regulation of cellular proliferation and differentiation. In mammalian cells Put is formed exclusively from L-ornithine by ornithine decarboxylase (ODC) and many potent ODC inhibitors are known. In bacteria, plants, and fungi Put is synthesized also from agmatine, which is formed from L-arginine by arginine decarboxylase (ADC). Here we demonstrate that the isosteric hydroxylamine analogue of agmatine (AO-Agm) is a new and very potent (IC50 3•10−8 M) inhibitor of E. coli ADC. It was almost two orders of magnitude less potent towards E. coli ODC. AO-Agm decreased polyamine pools and inhibited the growth of DU145 prostate cancer cells only at high concentration (1 mM). Growth inhibitory analysis of the Acremonium chrysogenum demonstrated that the wild type (WT) strain synthesized Put only from L-ornithine, while the cephalosporin C high-yielding strain, in which the polyamine pool is increased, could use both ODC and ADC to produce Put. Thus, AO-Agm is an important addition to the set of existing inhibitors of the enzymes of polyamine biosynthesis, and an important instrument for investigating polyamine biochemistry.

Regulation of polyamine biosynthesis by antizyme and some recent developments relating the induction of polyamine biosynthesis to cell growth

1985 ◽  
Vol 5 (3) ◽  
pp. 189-204 ◽  
Author(s):  
E. S. Canellakis ◽  
D. A. Kyriakidis ◽  
C. A. Rinehart ◽  
S.-C. Huang ◽  
C. Panagiotidis ◽  
...  
Keyword(s):  
Ornithine Decarboxylase ◽  
Mammalian Cells ◽  
Ribosomal Proteins ◽  
Physiological Role ◽  
Arginine Decarboxylase ◽  
Cell Of Origin ◽  
Polyamine Biosynthesis ◽  
Recent Developments

This review considers the role of antizyme, of amino acids and of protein synthesis in the regulation of polyamine biosynthesis.The ornithine decarboxylase of eukaryotic ceils and of Escherichia coli coli can be non-competitively inhibited by proteins, termed antizymes, which are induced by di-and poly- amines. Some antizymes have been purified to homogeneity and have been shown to be structurally unique to the cell of origin. Yet, the E. coli antizyme and the rat liver antizyme cross react and inhibit each other's biosynthetic decarboxylases. These results indicate that aspects of the control of polyamine biosynthesis have been highly conserved throughout evolution.Evidence for the physiological role of the antizyme in mammalian cells rests upon its identification in normal uninduced cells, upon the inverse relationship that exists between antizyme and ornithine decarboxylase as well as upon the existence of the complex of ornithine decarboxylase and antizyme in vivo. Furthermore, the antizyme has been shown to be highly specific; its Keq for ornithine decarboxylase is 1.4 × 1011 M-1. In addition, mammalian ceils contain an anti-antizyme, a protein that specifically binds to the antizyme of an ornithine decarboxylase-antizyme complex and liberates free ornithine decarboxylase from the complex. In B. coli, in which polyamine biosynthesis is mediated both by ornithine decarboxylase and by arginine decarboxylase, three proteins (one acidic and two basic) have been purified, each of which inhibits both these enzymes. They do not inhibit the biodegradative ornithine and arginine decarboxylases nor lysine decarboxylase. The two basic inhibitors have been shown to correspond to the ribosomal proteins S20/L26 and L34, respectively. The relationship of the acidic antizyme to other known B. coli proteins remains to be determined.

scholarly journals CSIG-29. STRUCTURAL AND FUNCTIONAL STUDIES OF PID1, A NOVEL GROWTH SUPPRESSOR IN BRAIN TUMORS

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi50-vi50
Author(s):  
Xiuhai Ren ◽  
Emi Hirsh ◽  
Min Mahdi ◽  
Fumiyaki Ito ◽  
Xiaojiang Chen ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Inhibitory Effect ◽  
Interaction Domain ◽  
Western Blots ◽  
E Coli ◽  
Functional Studies ◽  
Growth Inhibitory ◽  
Inhibitory Region ◽  
And Function ◽  
Growth Suppressor

Abstract BACKGROUND Brain cancers, including medulloblastomas and gliomas, are a major cause of death in children and adults. We reported that PID1 (Phosphotyrosine Interaction Domain containing 1) is a growth suppressor in medulloblastomas and gliomas (PMID: 24300787). PID1 also enhances the anti-tumor effects of chemotherapy (PMID: 28400607). We are now seeking to better understand the structure and mechanism of PID1, in order to utilize this knowledge to develop innovative PID1-based therapies. METHODS PID1 mutants were expressed in E. coli as MBP fusion proteins, purified and used for MALS analysis. Additional epitope tagged PID1 constructs (HA, V5, tGFP, eGFP) were generated, expressed in mammalian cells and analyzed by western blots, immunoprecipitation, and functional assays. RESULTS We carried out screening and testing of multiple mutants of purified PID1 and characterized them. Experiments in cell culture supported presence of similar findings in mammalian cells. Colony assays in glioma and medulloblastoma cell lines identified a region in PID1 that confers the most robust growth-inhibitory effect. Experiments are underway to further refine the boundaries and characteristics of this growth-inhibitory region. CONCLUSIONS This project, which is focused on better understanding of the structure and function of PID1, is uncovering novel aspects of its molecular function. Insights gained from this work will guide studies to develop innovative PID1-based therapies for gliomas and medulloblastomas.

scholarly journals Catalytic irreversible inhibition of bacterial and plant arginine decarboxylase activities by novel substrate and product analogues

1987 ◽  
Vol 242 (1) ◽  
pp. 69-74 ◽  
Author(s):  
A J Bitonti ◽  
P J Casara ◽  
P P McCann ◽  
P Bey
Keyword(s):  
Arginine Decarboxylase ◽  
Irreversible Inhibitor ◽  
Polyamine Biosynthesis ◽  
E Coli ◽  
First Order ◽  
Irreversible Inhibition ◽  
First Order Kinetics ◽  
Low Concentrations ◽  
Pseudo First Order

Arginine decarboxylase (ADC) activity from Escherichia coli and two plant species (oats and barley) was inhibited by five new substrate (arginine) and product (agmatine) analogues. The five compounds, (E)-alpha-monofluoromethyldehydroarginine (delta-MFMA), alpha-monofluoromethylarginine (MFMA), alpha-monofluoromethylagatine (FMA), alpha-ethynylagmatine (EA) and alpha-allenylagmatine (AA), were all more potent inhibitors of ADC activity than was alpha-difluoromethylarginine (DFMA), the only irreversible inhibitor of this enzyme described previously. The inhibition caused by the five compounds was apparently enzyme-activated and irreversible, since the loss of enzyme activity followed pseudo-first-order kinetics, was time-dependent, the natural substrate of ADC (arginine) blocked the effects of the inhibitors, and the inhibition remained after chromatography of inhibited ADC on Sephadex G-25 or on overnight dialysis of the enzyme. DFMA, FMA, delta-MFMA and MFMA were effective at very low concentrations (10 nM-10 microM) at inhibiting ADC activity in growing E. coli. FMA was also shown to deplete putrescine effectively in E. coli, particularly when combined with an inhibitor of ornithine decarboxylase, alpha-monofluoromethyl-putrescine. The potential uses of the compounds for the study of the role of polyamine biosynthesis in bacteria and plants is discussed.

Use of Uranium-Labeled Antibodies for Electron Microscopic Localization of Various Antigens in Mammalian Cells

1967 ◽  
Vol 25 ◽  
pp. 136-137
Author(s):  
J. P. Petrali ◽  
E. J. Donati ◽  
L. A. Sternberger
Keyword(s):  
Endoplasmic Reticulum ◽  
Hela Cells ◽  
Mammalian Cells ◽  
Specific Antiserum ◽  
Electron Microscopic ◽  
E Coli ◽  
Cytoplasmic Membranes ◽  
Viral Progeny ◽  
Ultrathin Sections ◽  
Electron Microscopic Localization

Specific contrast is conferred to subcellular antigen by applying purified antibodies, exhaustively labeled with uranium under immunospecific protection, to ultrathin sections. Use of Seligman’s principle of bridging osmium to metal via thiocarbohydrazide (TCH) intensifies specific contrast. Ultrathin sections of osmium-fixed materials were stained on the grid by application of 1) thiosemicarbazide (TSC), 2) unlabeled specific antiserum, 3) uranium-labeled anti-antibody and 4) TCH followed by reosmication. Antigens to be localized consisted of vaccinia antigen in infected HeLa cells, lysozyme in monocytes of patients with monocytic or monomyelocytic leukemia, and fibrinogen in the platelets of these leukemic patients. Control sections were stained with non-specific antiserum (E. coli).In the vaccinia-HeLa system, antigen was localized from 1 to 3 hours following infection, and was confined to degrading virus, the inner walls of numerous organelles, and other structures in cytoplasmic foci. Surrounding architecture and cellular mitochondria were unstained. 8 to 14 hours after infection, antigen was localized on the outer walls of the viral progeny, on cytoplasmic membranes, and free in the cytoplasm. Staining of endoplasmic reticulum was intense and focal early, and weak and diffuse late in infection.

Cell-Specific Chemical Delivery Using a Selective Nitroreductase–Nitroaryl Pair

2018 ◽  
Author(s):  
Todd D. Gruber ◽  
Chithra Krishnamurthy ◽  
Jonathan B. Grimm ◽  
Michael R. Tadross ◽  
Laura M. Wysocki ◽  
...  
Keyword(s):  
Small Molecules ◽  
Mammalian Cells ◽  
Target Cells ◽  
Specific Chemical ◽  
E Coli ◽  
Enzyme Substrate ◽  
Cell Specificity ◽  
General Chemical ◽  
Increased Activity ◽  
Enzyme Variant

<p>The utility of<b> </b>small molecules to probe or perturb biological systems is limited by the lack of cell-specificity. ‘Masking’ the activity of small molecules using a general chemical modification and ‘unmasking’ it only within target cells could overcome this limitation. To this end, we have developed a selective enzyme–substrate pair consisting of engineered variants of <i>E. coli</i> nitroreductase (NTR) and a 2‑nitro-<i>N</i>-methylimidazolyl (NM) masking group. To discover and optimize this NTR–NM system, we synthesized a series of fluorogenic substrates containing different nitroaromatic masking groups, confirmed their stability in cells, and identified the best substrate for NTR. We then engineered the enzyme for improved activity in mammalian cells, ultimately yielding an enzyme variant (enhanced NTR, or eNTR) that possesses up to 100-fold increased activity over wild-type NTR. These improved NTR enzymes combined with the optimal NM masking group enable rapid, selective unmasking of dyes, indicators, and drugs to genetically defined populations of cells.</p>

Occurrences of Giardia cysts in beach water

1998 ◽  
Vol 38 (12) ◽  
pp. 73-76 ◽  
Author(s):  
B. S. W. Ho ◽  
T.-Y Tam
Keyword(s):  
Health Risk ◽  
Water Sample ◽  
Water Samples ◽  
High Concentration ◽  
Potential Health ◽  
Infectious Dose ◽  
Potential Health Risk ◽  
E Coli ◽  
Beach Water ◽  
Giardia Cysts

A total of 64 beach water samples with various bacteriological quality (Grades 1 to 4) were analysed for their bacteriological and parasitological contents (E coli and Giardia cysts respectively). Results indicated that Giardia cysts were detected in less than 10% of the Grade 1 beach water samples with E coli concentrations of &lt;24/100mL. For Grades 2, 3 & 4 beach water samples, Giardia cysts were found, respectively, in 85, 50 and 64% of the samples. Except for one beach water sample which had an unusually high concentration of Giardia cysts (23 cysts/L), they were generally present at moderate concentrations (&lt;10 cysts/L) in all other beach water samples. Despite moderate levels of Giardia cysts present in beach water of different grades, the potential health risk faced by swimmers bathing in local beach water needs to be carefully assessed as Giardia is known to have a low infectious dose.

scholarly journals Use of Acellular Umbilical Cord-Derived Tissues in Corneal and Ocular Surface Diseases

Medicines ◽  
2021 ◽  
Vol 8 (2) ◽  
pp. 12
Author(s):  
Arianna A. Tovar ◽  
Ian A. White ◽  
Alfonso L. Sabater
Keyword(s):  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Ocular Surface ◽  
Cellular Proliferation ◽  
Mononuclear Cells ◽  
Autologous Serum ◽  
High Concentration ◽  
Ocular Surface Diseases ◽  
Natural Healing

Blood derived products have become a valuable source of tissue for the treatment of ocular surface diseases that are refractory to conventional treatments. These can be obtained from autologous or allogeneic sources (patient’s own blood or from healthy adult donors/umbilical cord blood, respectively). Allogeneic cord blood demonstrates practical advantages over alternatives and these advantages will be discussed herein. Umbilical cord blood (UCB) can be divided, generally speaking, into two distinct products: first, mononuclear cells, which can be used in regenerative ophthalmology, and second, the plasma/serum (an acellular fraction), which may be used in the form of eyedrops administered directly to the damaged ocular surface. The rationale for using umbilical cord serum (UCS) to treat ocular surface diseases such as severe dry eye syndrome (DES), persistent epithelial defects (PED), recurrent epithelial erosions, ocular chemical burns, graft versus host disease (GVHD), among others, is the considerably high concentration of growth factors and cytokines, mimicking the natural healing properties of human tears. Allogeneic serum also offers the opportunity for therapeutic treatment to patients who, due to poor heath, cannot provide autologous serum. The mechanism of action involves the stimulation of endogenous cellular proliferation, differentiation and maturation, which is highly efficient in promoting and enhancing corneal epithelial healing where other therapies have previously failed.

scholarly journals Nucleoside Analogues Are Potent Inducers of Pol V-mediated Mutagenesis

Biomolecules ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 843
Author(s):  
Balagra Kasim Sumabe ◽  
Synnøve Brandt Ræder ◽  
Lisa Marie Røst ◽  
Animesh Sharma ◽  
Eric S. Donkor ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Mutation Frequency ◽  
Nucleoside Analogues ◽  
Cancer Therapies ◽  
E Coli ◽  
Replicative Stress ◽  
Pol V ◽  
Dna And Rna ◽  
Sos Induction ◽  
Anti Cancer

Drugs targeting DNA and RNA in mammalian cells or viruses can also affect bacteria present in the host and thereby induce the bacterial SOS system. This has the potential to increase mutagenesis and the development of antimicrobial resistance (AMR). Here, we have examined nucleoside analogues (NAs) commonly used in anti-viral and anti-cancer therapies for potential effects on mutagenesis in Escherichia coli, using the rifampicin mutagenicity assay. To further explore the mode of action of the NAs, we applied E. coli deletion mutants, a peptide inhibiting Pol V (APIM-peptide) and metabolome and proteome analyses. Five out of the thirteen NAs examined, including three nucleoside reverse transcriptase inhibitors (NRTIs) and two anti-cancer drugs, increased the mutation frequency in E. coli by more than 25-fold at doses that were within reported plasma concentration range (Pl.CR), but that did not affect bacterial growth. We show that the SOS response is induced and that the increase in mutation frequency is mediated by the TLS polymerase Pol V. Quantitative mass spectrometry-based metabolite profiling did not reveal large changes in nucleoside phosphate or other central carbon metabolite pools, which suggests that the SOS induction is an effect of increased replicative stress. Our results suggest that NAs/NRTIs can contribute to the development of AMR and that drugs inhibiting Pol V can reverse this mutagenesis.

scholarly journals Overexpression of arginine decarboxylase in transgenic plants

1997 ◽  
Vol 325 (2) ◽  
pp. 331-337 ◽  
Author(s):  
Daniel BURTIN ◽  
Anthony J. MICHAEL
Keyword(s):  
Transgenic Plants ◽  
Arginine Decarboxylase ◽  
Polyamine Metabolism ◽  
Morphological Development ◽  
Polyamine Biosynthesis ◽  
Adenosylmethionine Decarboxylase ◽  
Two Generations ◽  
Key Enzyme ◽  
Polyamine Conjugate ◽  
And Control

The activity of arginine decarboxylase (ADC), a key enzyme in plant polyamine biosynthesis, was manipulated in two generations of transgenic tobacco plants. Second-generation transgenic plants overexpressing an oat ADC cDNA contained high levels of oat ADC transcript relative to tobacco ADC, possessed elevated ADC enzyme activity and accumulated 10–20-fold more agmatine, the direct product of ADC. In the presence of high levels of the precursor agmatine, no increase in the levels of the polyamines putrescine, spermidine and spermine was detected in the transgenic plants. Similarly, the activities of ornithine decarboxylase and S-adenosylmethionine decarboxylase were unchanged. No diversion of polyamine metabolism into the hydroxycinnamic acid–polyamine conjugate pool or into the tobacco alkaloid nicotine was detected. Activity of the catabolic enzyme diamine oxidase was the same in transgenic and control plants. The elevated ADC activity and agmatine production were subjected to a metabolic/physical block preventing increased, i.e. deregulated, polyamine accumulation. Overaccumulation of agmatine in the transgenic plants did not affect morphological development.

scholarly journals In Escherichia coli Ammonia Inhibits Cytochrome bo3 But Activates Cytochrome bd-I

Antioxidants ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 13
Author(s):  
Elena Forte ◽  
Sergey A. Siletsky ◽  
Vitaliy B. Borisov
Keyword(s):  
Escherichia Coli ◽  
Molecular Mechanisms ◽  
Dissociation Constants ◽  
Reductase Activity ◽  
Ammonia Concentration ◽  
High Concentration ◽  
E Coli ◽  
Cytochrome Bd ◽  
Cytochrome Bo3 ◽  
Oxygen Reductase

Interaction of two redox enzymes of Escherichia coli, cytochrome bo3 and cytochrome bd-I, with ammonium sulfate/ammonia at pH 7.0 and 8.3 was studied using high-resolution respirometry and absorption spectroscopy. At pH 7.0, the oxygen reductase activity of none of the enzymes is affected by the ligand. At pH 8.3, cytochrome bo3 is inhibited by the ligand, with 40% maximum inhibition at 100 mM (NH4)2SO4. In contrast, the activity of cytochrome bd-I at pH 8.3 increases with increasing the ligand concentration, the largest increase (140%) is observed at 100 mM (NH4)2SO4. In both cases, the effector molecule is apparently not NH4+ but NH3. The ligand induces changes in absorption spectra of both oxidized cytochromes at pH 8.3. The magnitude of these changes increases as ammonia concentration is increased, yielding apparent dissociation constants Kdapp of 24.3 ± 2.7 mM (NH4)2SO4 (4.9 ± 0.5 mM NH3) for the Soret region in cytochrome bo3, and 35.9 ± 7.1 and 24.6 ± 12.4 mM (NH4)2SO4 (7.2 ± 1.4 and 4.9 ± 2.5 mM NH3) for the Soret and visible regions, respectively, in cytochrome bd-I. Consistently, addition of (NH4)2SO4 to cells of the E. coli mutant containing cytochrome bd-I as the only terminal oxidase at pH 8.3 accelerates the O2 consumption rate, the highest one (140%) being at 27 mM (NH4)2SO4. We discuss possible molecular mechanisms and physiological significance of modulation of the enzymatic activities by ammonia present at high concentration in the intestines, a niche occupied by E. coli.

Sign in / Sign up

Export Citation Format

Share Document