scholarly journals An NAD+-dependent alanine dehydrogenase from a methylotrophic bacterium

1987 ◽  
Vol 244 (3) ◽  
pp. 565-570 ◽  
Author(s):  
E Bellion ◽  
F Tan
Keyword(s):  
Native Enzyme ◽  
Methylotrophic Bacterium ◽  
Subunit Structure ◽  
Pseudomonas Sp ◽  
Polyacrylamide Gels ◽  
Limited Extent ◽  
Ph Optimum ◽  
Alanine Dehydrogenase

A study was made of the NAD+-dependent alanine dehydrogenase (EC 1.4.1.1) elaborated by the methylotrophic bacterium Pseudomonas sp. strain MA when growing on succinate and NH4Cl. This enzyme was purified 400-fold and was found to be highly specific for NH3 and NAD+; however, hydroxypyruvate and bromopyruvate, but not alpha-oxoglutarate or glyoxylate, could replace pyruvate to a limited extent. The Mr of the native enzyme was shown to be 217,000, and electrophoresis in SDS/polyacrylamide gels revealed a minimum Mr of 53,000, suggesting a four-subunit structure. The enzyme, which has a pH optimum of 9.0, operated almost exclusively in the aminating direction in vitro. It was induced by NH3 or by alanine, and was repressed by growth on methylamine or glutamate. It is suggested that this enzyme has two roles in this organism, namely in NH3 assimilation and in alanine catabolism.

scholarly journals The nucleoside triphosphate–ribonucleic acid nucleotidyltransferase (EC 2.7.7.6) of Agrobacterium tumefaciens (Smith and Townsend) Conn. Purification and properties of the enzyme from the tumorigenic strain B6806

1974 ◽  
Vol 143 (3) ◽  
pp. 511-520 ◽  
Author(s):  
U. C. Knopf
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Gradient Centrifugation ◽  
Deae Cellulose ◽  
Nucleoside Triphosphate ◽  
Subunit Structure ◽  
Bacterial Cells ◽  
Protein Subunit ◽  
Ph Optimum ◽  
Protamine Sulphate

The RNA nucleotidyltransferase (RNA polymerase) of the plant-tumorigenic bacterium Agrobacterium tumefaciens was purified. The method involves the disruption of the bacterial cells with glass beads in a Waring Blendor, treatment with DEAE-cellulose, fractionation with (NH4)2SO4, protamine sulphate precipitation, DEAE-cellulose column chromatography and either glycerol-gradient centrifugation or phosphocellulose chromatography. The subunit structure of the highly purified enzyme is similar to, although not identical with, the RNA nucleotidyltransferase of Escherichia coli. It can be described as β′, β, χ1 and α (mol.wts. 160000, 150000, 98000, and 41000±10% respectively). χ1 is the temporary designation for a protein subunit, which might have the same functions as the σ subunit in E. coli. The enzyme of A. tumefaciens is rifampicin-sensitive, has a temperature optimum in vitro of 41±1°C and a pH optimum of 8.2±0.1. Mg2+ and Mn2+ are activators. The enzyme transcribes with different efficiencies artificial, viral, bacterial, plant and animal templates.

scholarly journals Pseudomonas sp. COW3 Produces New Bananamide-Type Cyclic Lipopeptides with Antimicrobial Activity against Pythium myriotylum and Pyricularia oryzae

Molecules ◽  
2019 ◽  
Vol 24 (22) ◽  
pp. 4170 ◽  
Author(s):  
Olumide Owolabi Omoboye ◽  
Niels Geudens ◽  
Matthieu Duban ◽  
Mickaël Chevalier ◽  
Christophe Flahaut ◽  
...  
Keyword(s):  
Antimicrobial Activity ◽  
Genome Mining ◽  
Antagonistic Activity ◽  
Pyricularia Oryzae ◽  
Pseudomonas Sp ◽  
Pythium Myriotylum ◽  
Cyclic Lipopeptides ◽  
Peptide Synthetase ◽  
Kendrick Mass Defect

Pseudomonas species are metabolically robust, with capacity to produce secondary metabolites including cyclic lipopeptides (CLPs). Herein we conducted a chemical analysis of a crude CLP extract from the cocoyam rhizosphere-derived biocontrol strain Pseudomonas sp. COW3. We performed in silico analyses on its whole genome, and conducted in vitro antagonistic assay using the strain and purified CLPs. Via LC-MS and NMR, we elucidated the structures of four novel members of the bananamide group, named bananamides D-G. Besides variability in fatty acid length, bananamides D-G differ from previously described bananamides A-C and MD-0066 by the presence of a serine and aspartic acid at position 6 and 2, respectively. In addition, bananamide G has valine instead of isoleucine at position 8. Kendrick mass defect (KMD) allowed the assignment of molecular formulae to bananamides D and E. We unraveled a non-ribosomal peptide synthetase cluster banA, banB and banC which encodes the novel bananamide derivatives. Furthermore, COW3 displayed antagonistic activity and mycophagy against Pythium myriotylum, while it mainly showed mycophagy on Pyricularia oryzae. Purified bananamides D-G inhibited the growth of P. myriotylum and P. oryzae and caused hyphal distortion. Our study shows the complementarity of chemical analyses and genome mining in the discovery and elucidation of novel CLPs. In addition, structurally diverse bananamides differ in their antimicrobial activity.

scholarly journals Plant holo-(acyl carrier protein) synthase

1988 ◽  
Vol 252 (1) ◽  
pp. 39-45 ◽  
Author(s):  
S A Elhussein ◽  
J A Miernyk ◽  
J B Ohlrogge
Keyword(s):  
Spinacia Oleracea ◽  
Acyl Carrier Protein ◽  
Ricinus Communis ◽  
Plant Cells ◽  
Gel Permeation Chromatography ◽  
Carrier Protein ◽  
Improved Method ◽  
Ph Optimum ◽  
Spinach Leaf

1. An improved method was developed for the assay of plant holo-(acyl carrier protein) synthase activity, using Escherichia coli acyl-(acyl carrier protein) synthetase as a coupling enzyme. 2. Holo-(acyl carrier protein) synthase was partially purified from spinach (Spinacia oleracea) leaves by a combination of (NH4)2SO4 fractionation and anion-exchange and gel-permeation chromatography. 3. The partially purified enzyme had a pH optimum of 8.2 and Km values of 2 microM, 72 microM and 3 mM for apo-(acyl carrier protein), CoA and Mg2+ respectively. Synthase activity was inhibited in vitro by the reaction product 3′,5′-ADP. 4. Results from the fractionation of spinach leaf and developing castor-oil-seed (Ricinus communis) endosperm cells were consistent with a cytosolic localization of holo-(acyl carrier protein) synthase activity in plant cells.

scholarly journals Engineering a bioluminescent indicator for cyclic AMP-dependent protein kinase

1991 ◽  
Vol 279 (3) ◽  
pp. 727-732 ◽  
Author(s):  
G B Sala-Newby ◽  
A K Campbell
Keyword(s):  
Protein Kinase ◽  
Cyclic Amp ◽  
Specific Activity ◽  
Phosphorylation Site ◽  
Live Cells ◽  
Phosphorylation Sites ◽  
Dependent Protein Kinase ◽  
Ph Optimum ◽  
Dependent Protein

cDNA coding for the luciferase in the firefly Photinus pyralis was amplified in vitro to generate cyclic AMP-dependent protein kinase phosphorylation sites. The DNA was transcribed and translated to generate light-emitting protein. A valine at position 217 was mutated to arginine to generate a site RRFS and the heptapeptide kemptide, the phosphorylation site of the porcine pyruvate kinase, was added at the N- or C-terminus of the luciferase. The proteins carrying phosphorylation sites were characterized for their specific activity, pI, effect of pH on the colour of the light emitted and effect of the catalytic subunit of protein kinase A in the presence of ATP. Only one of the recombinant proteins (RRFS) was significantly different from wild-type luciferase. The RRFS mutant had a lower specific activity, lower pH optimum, emitted greener light at low pH and when phosphorylated it decreased its activity by up to 80%. This latter effect was reversed by phosphatase. This recombinant protein is a good candidate to measure for the first time cyclic AMP-dependent phosphorylation in live cells.

scholarly journals Activity of extracellular compounds of Pseudomonas sp. against Xanthomonas axonopodis in vitro and bacterial leaf blight in eucalyptus

2012 ◽  
Vol 37 (4) ◽  
pp. 233-238 ◽  
Author(s):  
Lucilene P. Lopes ◽  
Admiltom G. Oliveira Jr ◽  
Jamile P.O. Beranger ◽  
Cíntia G. Góis ◽  
Fernanda C.S. Vasconcellos ◽  
...  
Keyword(s):  
Leaf Blight ◽  
Bacterial Leaf Blight ◽  
Pseudomonas Sp ◽  
Xanthomonas Axonopodis

scholarly journals Saprophytic microorganisms with potential for biological control of Botrytis cinerea on Geraldton waxflower flowers

2001 ◽  
Vol 41 (5) ◽  
pp. 697 ◽  
Author(s):  
D. R. Beasley ◽  
D. C. Joyce ◽  
L. M. Coates ◽  
A. H. Wearing
Keyword(s):  
Botrytis Cinerea ◽  
Mycelial Growth ◽  
Germ Tube ◽  
Conidial Germination ◽  
Pseudomonas Sp ◽  
Trichoderma Spp ◽  
Fusarium Sp ◽  
Flower Abscission ◽  
Germ Tube Elongation

Saprophytic bacteria, yeasts and filamentous fungi were isolated from Geraldton waxflower flowers and screened to identify potential antagonism towards Botrytis cinerea. Isolates from other sources (e.g. avocado) were also tested. Isolates were initially screened in vitro for inhibition of B. cinerea conidial germination, germ tube elongation and mycelial growth. The most antagonistic bacteria, yeasts and fungi were selected for further testing on detached waxflower flowers. Conidia of the pathogen were mixed with conidia or cells of the selected antagonists, co-inoculated onto waxflower flowers, and the flowers were sealed in glass jars and incubated at 20˚C. The number of days required for the pathogen to cause flower abscission was determined. The most antagonistic bacterial isolate, Pseudomonas sp. 677, significantly reduced conidial germination and retarded germ tube elongation of B. cinerea. None of the yeast or fungal isolates tested was found to significantly reduce conidial germination or retard germ tube elongation, but several significantly inhibited growth of B. cinerea. Fusarium sp., Epicoccum sp. and Trichoderma spp. were the most antagonistic of these isolates. Of the isolates tested on waxflower, Pseudomonas sp. 677 was highly antagonistic towards B. cinerea and delayed waxflower abscission by about 3 days. Trichoderma harzianum also significantly delayed flower abscission. However, as with most of the fungal antagonists used, inoculation of waxflower flowers with this isolate resulted in unsightly mycelial growth.

scholarly journals Exploitation of endophytic Pseudomonas sp. for plant growth promotion and colonization in rice

2017 ◽  
Vol 9 (3) ◽  
pp. 1310-1316
Author(s):  
Gurjot Kaur ◽  
Poonam Sharma ◽  
Deepika Chhabra ◽  
Kailash Chand ◽  
Gurjit Singh Mangat
Keyword(s):  
Plant Growth ◽  
Plant Growth Promotion ◽  
Growth Promotion ◽  
Acc Deaminase ◽  
Pseudomonas Sp ◽  
Bacterial Isolates ◽  
Pgp Traits ◽  
P Solubilization ◽  
Fluorescent Pigment

The present investigation was carried out to exploit bacterial endophytes associated with root and leaf tissue of rice plant for plant growth promotion (PGP) and colonization study in vitro. Total 10 endophytic bacterial isolates (Pseudomonas sp.) were evaluate for PGP traits like P solubilization, production of Indole acetic acid (IAA), siderophore, ACC deaminase, protease, cellulase, fluorescent pigment, urease and denitrification activity. Out of 10 endophytic bacteria 30 %, 60 %, 20 %, 70 %, 10 % and 10 % were positive for siderophore, protease, cellulase, fluorescent pigment, urease and denitrification respectively. Maximum IAA production was recorded with isolate LRBLE7 (18.8 μgml-1) followed by LRBRE4 (16.0 μgml-1) and maximum P-solubilization was recorded with isolate LRBRE4 (5.8 mg 100 ml-1) followed by LRBLE7 (4.4 mg 100 ml-1). ACC deaminase production was recorded with isolate LRBLE6 (O.D=0.352 nm) followed by LRBRE5 (O.D=0.324nm). Three potential isolates (LRBRE4, LRBRE6 and LRBLE7) were selected on the basis of multiple PGP traits and were subjected to colonization study of rice seedling in vitro. Potential bacterial isolates can be exploited for improving growth and productivity in rice under sustainable management system.

An Electron-Microscope Study of the in vitro Transformation of Human Leucocytes

1967 ◽  
Vol 2 (3) ◽  
pp. 359-370
Author(s):  
J. A. CHAPMAN ◽  
M. W. ELVES ◽  
J. GOUGH
Keyword(s):  
Endoplasmic Reticulum ◽  
Electron Microscope ◽  
Messenger Rna ◽  
Human Peripheral Blood ◽  
Limited Extent ◽  
Single Particles ◽  
Blast Cells ◽  
Protein Secreting

Electron-microscope studies of cultured small lymphocytes from human peripheral blood transforming into larger blastoid cells in the presence of phytohaemagglutinin (PHA) show that the transformed cell possesses the preliminary stages of development of a protein-synthesizing system. The transformed blastoid cell has abundant ribosomes, although, in contrast with in vivo protein-secreting cells, many of these occur as single particles with only a small proportion Linked in polysomal clusters. Endoplasmic reticulum membranes occur to a very limited extent and with a marked paucity of attached ribosomal particles; the few attached particles are usually located in groups. Some endoplasmic reticulum membranes revealed degenerative changes in otherwise normal cells. A moderately well-developed Golgi apparatus was a characteristic feature of the cells. Apart from the relatively low proportion of polysomes, in vitro PHA-transformed blastoid cells are identical in fine structure to in vivo blast cells (otherwise known as immunoblasts, haemocytoblasts, etc.) occurring in the immune response. It is suggested that messenger-RNA production in PHA-stimulated transformed cells may be reduced and that this could explain the limited number of polysomes and the restricted development of the endoplasmic reticulum.

Requirements for extracellular metabolism of pulmonary surfactant: tentative identification of serine protease

1992 ◽  
Vol 262 (4) ◽  
pp. L446-L453 ◽  
Author(s):  
N. J. Gross ◽  
R. M. Schultz
Keyword(s):  
Serine Protease ◽  
Surface Area ◽  
Pulmonary Surfactant ◽  
Lamellar Body ◽  
Alveolar Epithelium ◽  
Alveolar Type ◽  
Type Ii Cells ◽  
Type Ii ◽  
Ph Optimum

Pulmonary alveolar surfactant is secreted by the alveolar epithelium in the form of lamellar bodylike structures that evolve sequentially into tubular myelin and vesicular forms that can be separated by centrifugation. Using an in vitro procedure by which the extracellular metabolism of pulmonary surfactant can be mimicked, namely cyclic variation in surface area, we previously reported that serine protease activity, which we called “convertase,” was required for the conversion of tubular myelin to the vesicular form. In the present studies we explored the biochemical requirements of this activity and sought the enzyme in alveolar products. Convertase activity has unusual requirements; in addition to being dependent on repetitive variations in surface area (cycling), it requires the presence of a high g fraction of lung secretions that is heat stable and not inhibitable by diisopropyl fluorophosphate (DFP) or alpha 1-antitrypsin, both typical serine protease inhibitors. The enzyme does not require calcium ions and has a pH optimum of 7.4. Convertase appears to be a component of surfactant itself because the ability of purified surfactant to convert to the vesicular form on cycling is impaired by pretreating it with DFP. A protein of Mr 75,000 that reacts with DFP and is heat sensitive was found in alveolar lavage, lamellar body preparations, and lung homogenate. It copurifies with lung surfactant in sucrose gradients. A similar DFP-reactive protein was observed in stable human neoplastic peripheral airway cell lines that express type II properties, suggesting that it may be a product of type II cells. We tentatively conclude that surfactant convertase is a 75,000 serine protease that is closely associated with surfactant phospholipid and that may be a product of alveolar type II cells.

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