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Purification and characterization of a phospholipase by Photobacterium damselae subsp. piscicida from cobia Rachycentron canadum.
- Source :
-
Journal of basic microbiology [J Basic Microbiol] 2014 Sep; Vol. 54 (9), pp. 969-75. Date of Electronic Publication: 2013 Jun 21. - Publication Year :
- 2014
-
Abstract
- Toxicity of the extracellular products (ECPs) and the lethal attributes of phospholipase secreted by pathogenic Photobacterium damselae subsp. piscicida from cobia Rachycentron canadum was studied. An extracellular lethal toxin in the ECPs was partially purified by using Fast Protein Liquid Chromatography system. A protein band (27 kDa) exhibited phospholipase activity on Native-PAGE (by 0.3% egg yolk agar-overlay), was excised and eluted. The pI value of the purified phospholipase was determined as 3.65 and was determined as a phospholipase C by using the Amplex™ Red phosphatidylcholine -Specific phospholipase C Assay kit. The phospholipase showed maximum activity at temperature around 4-40 °C and maximal activity at pH between 8 and 9. The enzyme was inhibited by ethylenediamine-tetraacetic acid (EDTA) and sodium dodecyl sulfate (SDS); but was activated by Ca(2+) and Mg(2+) and inactivated by Zn(2+) and Cu(2+) . Both the ECPs and phospholipase were hemolytic against erythrocytes of cobia and lethal to the fish with LD50 values of 3.25 and 0.91 µg protein g(-1) fish, respectively. In toxicity neutralization test, the rabbit antisera against the phospholipase could neutralize the toxicity of ECPs, indicating that the phospholipase is a major extracellular toxin produced by the bacterium.<br /> (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)
- Subjects :
- Animals
Chromatography, Liquid
Electrophoresis, Polyacrylamide Gel
Enzyme Activators analysis
Enzyme Inhibitors analysis
Enzyme Stability
Hemolysin Proteins chemistry
Hemolysin Proteins toxicity
Hemolysis
Hydrogen-Ion Concentration
Isoelectric Point
Lethal Dose 50
Molecular Weight
Temperature
Type C Phospholipases chemistry
Type C Phospholipases toxicity
Fishes microbiology
Hemolysin Proteins isolation & purification
Hemolysin Proteins metabolism
Photobacterium enzymology
Type C Phospholipases isolation & purification
Type C Phospholipases metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1521-4028
- Volume :
- 54
- Issue :
- 9
- Database :
- MEDLINE
- Journal :
- Journal of basic microbiology
- Publication Type :
- Academic Journal
- Accession number :
- 23787821
- Full Text :
- https://doi.org/10.1002/jobm.201200764