scholarly journals Purification and characterization of citrate synthase isoenzymes from Pseudomonas aeruginosa

1995 ◽  
Vol 309 (2) ◽  
pp. 507-511 ◽  
Author(s):  
C G Mitchell ◽  
S C K Anderson ◽  
E M T el-Mansi
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Citrate Synthase ◽  
Electrophoretic Analysis ◽  
Purification And Characterization ◽  
Sds Page ◽  
Dimethyl Suberimidate ◽  
Regulatory Properties ◽  
Single Subunit ◽  
Chemical Cross Linking

Two types of citrate synthase (CS) have been purified from Pseudomonas aeruginosa, a ‘large’ form (CSI) and a ‘small’ form (CSII). The M(r)s of the CSI and CSII isoenzymes were determined to be 240,000 +/- 16,000 (mean +/- S.E.M.) and 80,300 +/- 3800 respectively. Chemical cross-linking of the native enzymes with either dimethyl suberimidate or glutaraldehyde followed by electrophoretic analysis by SDS/PAGE showed that CSI is a hexamer and CSII is a dimer. SDS/PAGE showed that CSI and CSII each consist of a single subunit type, of M(r) 42,000 +/- 2000 and M(r) 36,500 +/- 2000 respectively. CSI and CSII were also shown to be distinct kinetically, immunologically and in terms of their regulatory properties. It is suggested that the CS isoenzymes are products of different structural genes.

Purification and characterization of an exochitinase from Bacillus thuringiensis subsp. aizawai and its action against phytopathogenic fungi

2006 ◽  
Vol 52 (7) ◽  
pp. 651-657 ◽  
Author(s):  
Luis Morales de la Vega ◽  
J Eleazar Barboza-Corona ◽  
Maria G Aguilar-Uscanga ◽  
Mario Ramírez-Lepe
Keyword(s):  
Bacillus Thuringiensis ◽  
Sodium Dodecyl ◽  
Sclerotium Rolfsii ◽  
Phytopathogenic Fungi ◽  
Purification And Characterization ◽  
Chitinolytic Enzyme ◽  
Bacillus Thuringiensis Subsp ◽  
Sds Page ◽  
Fusarium Sp

A chitinolytic enzyme from Bacillus thuringiensis subsp. aizawai has been purified and its molecular mass was estimated ca. 66 kDa by sodium dodecyl sulfate – polyacryamide gel electrophoresis (SDS–PAGE). The enzyme was able to hydrolyze chitin to chitobiosides but not carboxymethylcellulose, cellulose, pullulan, and laminarin. Optimal pH and temperature were detected at 6 and 50 °C, respectively. Stability, in the absence of substrate, was observed at temperatures less than 60 °C and pH between 5 and 8. Enzyme activity was significantly inhibited by K+ and EDTA and completely inhibited by Hg2+. Purified chitinase showed lytic activity against cell walls from six phytopathogenic fungi and inhibited the mycelial growth of both Fusarium sp. and Sclerotium rolfsii. The biocontrol efficacy of the enzyme was tested in the protection of bean seeds infested with six phytopathogenic fungi.Key words: chitinase, Bacillus thuringiensis, purification, phytopathogenic fungi.

scholarly journals Purification and Characterization of Pseudomonas aeruginosa PAO1 Asparaginase

2015 ◽  
Vol 28 ◽  
pp. 72-77 ◽  
Author(s):  
Takeshi Kuwabara ◽  
Asep A. Prihanto ◽  
Mamoru Wakayama ◽  
Kazuyoshi Takagi
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Purification And Characterization ◽  
Pseudomonas Aeruginosa Pao1

Purification and Characterization of Cytochrome c6 from the Unicellular Green Alga Scenedesmus obliquus

1997 ◽  
Vol 52 (11-12) ◽  
pp. 740-746 ◽  
Author(s):  
Röbbe Wünschiers ◽  
Thomas Zinn ◽  
Dietmar Linder ◽  
Rüdiger Schulz
Keyword(s):  
Cytochrome C ◽  
Green Alga ◽  
Scenedesmus Obliquus ◽  
Terminal Sequence ◽  
Ammonium Sulfate Precipitation ◽  
Purification And Characterization ◽  
Unicellular Green Alga ◽  
Sds Page ◽  
Monoraphidium Braunii

Abstract Purification of a soluble cytochrome c6 from the unicellular green alga Scenedesmus obliquus by a simple and rapid method is described. The purification procedure includes ammonium sulfate precipitation and non-denaturating PAGE. The N-terminal sequence of the first 20 amino acids was determined and shows 85% similarity and 75% identity to the sequence of cytochrome c6 from the green alga Monoraphidium braunii. The ferrocyto-chrome shows typical UV/VIS absorption peaks at 552.9, 521.9 and 415.7 nm. The apparent molecular mass was estimated to be 12 kD a by SDS-PAGE. EPR-spectroscopy at 20K shows resonances indicative for two distinct low-spin heme forms.

Purification and characterization of phosphotriesterases from Pseudomonas aeruginosa F10B and Clavibacter michiganense subsp. insidiosum SBL11

2006 ◽  
Vol 52 (2) ◽  
pp. 157-168 ◽  
Author(s):  
Subhas Das ◽  
Dileep Kumar Singh
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Molecular Mass ◽  
Half Life ◽  
Indoleacetic Acid ◽  
Bacterial Strains ◽  
Purification And Characterization ◽  
Temperature Optimum ◽  
Microbial Biodegradation ◽  
Membrane Bound

A microbial biodegradation of monocrotophos was studied in the present investigation. The monocrotophos-degrading enzyme was purified and characterized from two soil bacterial strains. The cells were disrupted and the membrane-bound fractions were studied for purification and characterization. Solubilization of the membrane-bound fractions released nearly 80% of the bound protein. Phase separation further enriched the enzyme fraction 34–41 times. The enzyme phosphotriesterase (PTE) from both the strains was purified to more than 1000-fold with 13%–16% yield. Purified PTE from Clavibacter michiganense subsp. insidiosum SBL11 is a monomeric enzyme with a molecular mass of 43.5 kDa (pI of 7.5), while PTE from Pseudomonas aeruginosa F10B is a heterodimeric enzyme with a molecular mass of 43 and 41 kDa (pI of 7.9 and 7.35). Both purified enzymes are stable enzymes with peak activity at pH 9.0. The enzyme from strain F10B was more thermostable (half-life = 7.3 h) than that from SBL11 (half-life = 6.4 h at 50 °C), while both showed the same temperature optimum of 37 °C. Inhibitors like dithiothreitol and EDTA inhibited the purified enzyme, while p-chloromercuribenzoic acid and indoleacetic acid had a very little effect.Key words: biodegradation, monocrotophos, phosphotriesterase, Pseudomonas aeruginosa F10B, Clavibacter michiganense subsp. insidiosum SBL11.

scholarly journals Purification and characterization of pig kidney aminopeptidase P. A glycosyl-phosphatidylinositol-anchored ectoenzyme

1990 ◽  
Vol 267 (2) ◽  
pp. 509-515 ◽  
Author(s):  
N M Hooper ◽  
J Hryszko ◽  
A J Turner
Keyword(s):  
Chelating Agents ◽  
Inhibitory Effect ◽  
Purification And Characterization ◽  
Glycosyl Phosphatidylinositol ◽  
Pig Kidney ◽  
Sds Page ◽  
Cyclic Phosphate ◽  
Aminopeptidase P ◽  
Mild Acid

Aminopeptidase P (EC 3.4.11.9) was solubilized from pig kidney membranes with bacterial phosphatidylinositol-specific phospholipase C (PI-PLC) and then purified by a combination of anion-exchange and hydrophobic-interaction chromatographies. Contaminating peptidase activities were removed by selective affinity chromatography. The purified enzyme was apparently homogeneous on SDS/PAGE with an Mr of 91,000. Enzymic deglycosylation revealed that aminopeptidase P is a glycoprotein, with up to 25% by weight of the protein being due to the presence of N-linked sugars. The phospholipase-solubilized aminopeptidase P was recognized by an antiserum to the cross-reacting determinant (CRD) characteristic of the glycosyl-phosphatidylinositol anchor. This recognition was abolished by mild acid treatment or deamination with HNO2, indicating that the CRD was due exclusively to the inositol 1,2-cyclic phosphate ring epitope generated by the action of PI-PLC. The activity of aminopeptidase P was inhibited by chelating agents and was stimulated by Mn2+ or Co2+ ions, confirming the metallo-enzyme nature of this peptidase. Selective inhibitors of other aminopeptidases (actinonin, amastatin, bestatin and puromycin) had little or no inhibitory effect.

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