scholarly journals The bacterial oxidation of N-methylisonicotinate, a photolytic product of Paraquat

1972 ◽  
Vol 127 (5) ◽  
pp. 833-844 ◽  
Author(s):  
C. G. Orpin ◽  
M. Knight ◽  
W. C. Evans
Keyword(s):  
Enzyme System ◽  
Soluble Enzyme ◽  
Bacterial Oxidation ◽  
Gram Negative ◽  
Whole Cells ◽  
Photodegradation Product ◽  
Enzyme Systems ◽  
A Cell ◽  
Methyl Derivatives ◽  
Bacterium Strain

Two bacteria have been isolated that are capable of oxidizing N-methylisonicotinate, a photodegradation product of Paraquat (1.1′-dimethyl-4,4′-bipyridylium ion). N-Methylisonicotinate-grown cells of strain 4C1, a Gram-positive rod, oxidized 2-hydroxy-N-methylisonicotinate without lag. Cell-free extracts of these cells converted 2-hydroxyisonicotinate into 2,6-dihydroxyisonicotinate; the reaction did not require molecular oxygen. Maleamate was deamidated and maleate isomerized to fumarate by soluble enzyme systems. [14C]Formaldehyde was isolated as the dimedone derivative from the supernatant of a cell suspension oxidizing N-[14C]methylisonicotinate, and no [14C]-methylamine was detected. Whole cells incubated with N-methyl[carboxy-14C]isonicotinate released 95% of the radioactivity as 14CO2. The second bacterium, strain 4C2, a Gram-negative rod, did not oxidize any of the mono- or di-hydroxypyridines or their N-methyl derivatives that were available or could be synthesized; nor did cell-free extracts oxidize any of these compounds. Methylamine was oxidized by whole cells without lag; cell-free extracts converted methylamine into formaldehyde when a soluble enzyme system requiring an electron acceptor was used; formaldehyde was oxidized to formate and formate to CO2 by enzyme systems requiring NAD+.

scholarly journals The bacterial oxidation of picolinamide, a photolytic product of Diquat

1972 ◽  
Vol 127 (5) ◽  
pp. 819-831 ◽  
Author(s):  
C. G. Orpin ◽  
M. Knight ◽  
W. C. Evans
Keyword(s):  
Molecular Oxygen ◽  
Ultrasonic Treatment ◽  
Substrate Concentration ◽  
Sodium Arsenite ◽  
Culture Media ◽  
High Substrate Concentration ◽  
Bacterial Oxidation ◽  
Gram Negative ◽  
Whole Cells ◽  
Culture Supernatants

The pathway of oxidation of picolinamide (pyridine-2-carboxamide) by a Gram-negative rod has been elucidated. Under conditions of high pH, restricted aeration and high substrate concentration, whole cells released 2,5-dihydroxypyridine into culture supernatants. Sodium arsenite at 5mm caused whole cells to accumulate 6-hydroxypicolinate, and, at 1mm, pyruvate, in culture media. Whole cells oxidized picolinamide, picolinate, 6-hydroxypicolinate, maleamate and maleate without lag. Cell-free extracts converted picolinamide into picolinate, and hydroxylated picolinate to 6-hydroxypicolinate. The hydroxylase was particulate, but could be solubilized by ultrasonic treatment; it required NAD+ for activity, and did not require molecular oxygen. 2,5-Dihydroxypyridine was converted into maleamate and formate by an oxygenase requiring GSH and Fe2+. Maleamate was deamidated to maleate, and maleate isomerized to fumarate, by unsupplemented extracts.

scholarly journals Preparation of some immobilized linked enzyme systems and their use in the automated determination of disaccharides

1974 ◽  
Vol 137 (1) ◽  
pp. 25-32 ◽  
Author(s):  
D. J. Inman ◽  
W. E. Hornby
Keyword(s):  
Glucose Oxidase ◽  
Enzyme System ◽  
Inside Surface ◽  
Enzyme Systems ◽  
Nylon Tube ◽  
In Series ◽  
Automated Determination

1. Glucose oxidase (EC 1.1.3.4), amyloglucosidase (EC 3.2.1.3), invertase (EC 3.2.1.26) and β-galactosidase (EC 3.2.1.23) were covalently attached via glutaraldehyde to the inside surface of nylon tube. 2. The linked enzyme system, comprising invertase immobilized within a nylon tube acting in series with glucose oxidase immobilized in a similar way, was used for the automated determination of sucrose. 3. The linked enzyme system, comprising β-galactosidase immobilized within a nylon tube acting in series with glucose oxidase immobilized in a similar way, was used for the automated determination of lactose. 4. The linked enzyme system, comprising amyloglucosidase immobilized within a nylon tube acting in series with glucose oxidase immobilized in a similar way, was used for the automated determination of maltose. 5. Mixtures of glucose oxidase and amyloglucosidase were immobilized within the same piece of nylon tube and used for the automated determination of maltose. 6. Mixtures of glucose oxidase and invertase were immobilized within the same piece of nylon tube and used for the automated determination of sucrose.

Formation of the nitrogen-fixing enzyme system in Azotobacter vinelandii

1968 ◽  
Vol 14 (1) ◽  
pp. 25-31 ◽  
Author(s):  
G. W. Strandberg ◽  
P. W. Wilson
Keyword(s):  
Color Change ◽  
Azotobacter Vinelandii ◽  
Nitrogenase Activity ◽  
Color Difference ◽  
Potassium Nitrate ◽  
Whole Cells ◽  
System A ◽  
Growth System ◽  
A Cell ◽  
Enzyme Formation

The formation and activity of nitrogenase2 in Azotobacter vinelandii OP was examined using a cell-free assay system. A lag period of about 30 min occurred between the exhaustion of the combined nitrogen source and growth on N2. Cells grown on ammonium acetate or potassium nitrate had no detectable nitrogenase activity. Nitrogenase activity appeared in cells, grown under a flowing gas phase of 20% O2 – 60% He, about 45 min after the exhaustion of ammonia. Nitrogenase formation was inhibited in a closed system with an atmosphere containing 40% O2 but not by one containing 20% O2. Hydrogen did not inhibit enzyme formation. The question of whether N2 is required for the formation of the enzyme could not be answered as this gas could not be completely eliminated from the growth system. Chloramphenicol prevented the formation of the enzyme and inhibited nitrogen fixation in whole cells, but had no effect on cell-free enzyme activity. A brief rise in turbidity which occurred during nitrogenase formation appeared to be due to a color change in the cells from reddish brown to dark brown. Spectrophotometric examination of extracts from ammonia- and N2-grown cells did not reveal any components responsible for this color difference, but this result may reflect only the presence of interfering substances in the crude extract.

scholarly journals Cucurbitacins are insect steroid hormone antagonists acting at the ecdysteroid receptor

1997 ◽  
Vol 327 (3) ◽  
pp. 643-650 ◽  
Author(s):  
Laurence DINAN ◽  
Pensri WHITING ◽  
Jean-Pierre GIRAULT ◽  
René LAFONT ◽  
S. Tarlochan DHADIALLA ◽  
...  
Keyword(s):  
Morphological Changes ◽  
Antagonistic Activity ◽  
Side Chain ◽  
Ecdysteroid Receptor ◽  
Whole Cells ◽  
Definitive Evidence ◽  
Permanent Cell ◽  
A Cell ◽  
Gel Shift Assay ◽  
Hormone Antagonists

Two triterpenoids, cucurbitacins B and D, have been isolated from seeds of Iberis umbellata (Cruciferae) and shown to be responsible for the antagonistic activity of a methanolic extract of this species in preventing the 20-hydroxyecdysone (20E)-induced morphological changes in the Drosophila melanogaster BII permanent cell line. With a 20E concentration of 50 nM, cucurbitacins B and D give 50% responses at 1.5 and 10 μM respectively. Both cucurbitacins are able to displace specifically bound radiolabelled 25-deoxy-20-hydroxyecdysone (ponasterone A) from a cell-free preparation of the BII cells containing ecdysteroid receptors. The Kd values for cucurbitacins B and D (5 and 50 μM respectively) are similar to the concentrations required to antagonize 20E activity with whole cells. Cucurbitacin B (cucB) prevents stimulation by 20E of an ecdysteroid-responsive reporter gene in a transfection assay. CucB also prevents the formation of the Drosophila ecdysteroid receptor/Ultraspiracle/20E complex with the hsp27 ecdysteroid response element as demonstrated by gel-shift assay. This is therefore the first definitive evidence for the existence of antagonists acting at the ecdysteroid receptor. Preliminary structure/activity studies indicate the importance of the Δ23-22-oxo functional grouping in the side chain for antagonistic activity. Hexanorcucurbitacin D, which lacks carbon atoms C-22 to C-27, is found to be a weak agonist rather than an antagonist. Moreover, the side chain analogue 5-methylhex-3-en-2-one possesses weak antagonistic activity.

Select Gram-negative Aerobic Bacteria

2012 ◽  
pp. 90-101
Author(s):  
David R. McNamara ◽  
Franklin R. Cockerill
Keyword(s):  
Cell Wall ◽  
Sepsis Syndrome ◽  
Vital Role ◽  
Klebsiella Pneumonia ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Neisseria Meningitides ◽  
A Cell ◽  
Normal Human ◽  
Specific Species

Gram-negative bacteria may be rod-shaped (bacilli), spherical (cocci), oval, helical, or filamentous. Cytoplasmic membrane is surrounded by a cell wall consisting of a peptidoglycan layer and an outer cell membrane. Gram-negative bacteria are widely distributed in the natural environment. They are commensals with many animals and play a vital role in normal human physiology as intestinal commensals. Gram-negative bacteria are the cause of various human illnesses. The gram-negative bacterial cell wall contains various lipopolysaccharide endotoxins. Endotoxins trigger intense inflammation and the sepsis syndrome during infection. Specific species of gram-negative bacteria such as Neisseria meningitides, Moraxella catarrhalis, Acinetobacter, Vibrio, Klebsiella pneumonia, Salmonella, Pseudomonas aeruginosa, and Haemophilus influenza are reviewed.

Functional analysis of the lysis genes of Staphylococcus aureus phage P68 in Escherichia coli

Microbiology ◽  
2005 ◽  
Vol 151 (7) ◽  
pp. 2331-2342 ◽  
Author(s):  
Marian Takáč ◽  
Angela Witte ◽  
Udo Bläsi
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Functional Analysis ◽  
Transmembrane Domains ◽  
Class I ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Reading Frame ◽  
Double Stranded Dna ◽  
A Cell

Double-stranded DNA phages of both Gram-positive and Gram-negative bacteria typically use a holin–endolysin system to achieve lysis of their host. In this study, the lysis genes of Staphylococcus aureus phage P68 were characterized. P68 gene lys16 was shown to encode a cell-wall-degrading enzyme, which causes cell lysis when externally added to clinical isolates of S. aureus. Another gene, hol15, was identified embedded in the −1 reading frame at the 3′ end of lys16. The deduced Hol15 protein has three putative transmembrane domains, and thus resembles class I holins. An additional candidate holin gene, hol12, was found downstream of the endolysin gene lys16 based on two predicted transmembrane domains of the encoded protein, which is a typical trait of class II holins. The synthesis of either Hol12 or Hol15 resulted in growth retardation of Escherichia coli, and both hol15 and hol12 were able to complement a phage λ Sam mutation. The hol15 gene has a dual start motif beginning with the codons Met1-Lys2-Met3…. Evidence is presented that the hol15 gene encodes a lysis inhibitor (anti-holin) and a lysis effector (actual holin). As depolarization of the membrane converted the anti-holin to a functional holin, these studies suggested that hol15 functions as a typical dual start motif class I holin. The unusual arrangement of the P68 lysis genes is discussed.

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