Facile isolation of endo-pectate lyase from Erwinia carotovora based on electrostatic interaction.
Endo-pectate lyase (PATE) from Erwinia carotovora was selectively cosedimented with extracellularly produced lipopolysaccharide-lipid complex (LPSLC) through dialysis of the cell free culture broth. The selective isolation of PATE was confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The cosedimentation of the PATE with LPSLC was initiated by decreasing conductivity of the solution and terminated at approx 1 m siemens (mScm-1). As much as 62% of PATE activity in the culture broth was removed by precipitation. PATE was isolated from the precipitate by gel chromatography. The cosedimentation of PATE with LPSLC was remarkably affected by pH or ionic strength. The addition of polycationic peptide polymyxin B sulfate or a metal chloride affected the interaction. The cosedimentation was diminished by acetylation of the free amino groups of PATE. From these results, it was confirmed that the cosedimentation was induced by electrostatic interaction.
Fukuoka
S
S
Government Industrial Research Institute, Takamatsu, Japan.
Kamishima
H
H
Sode
K
K
Karube
I
I
D000107:Acetylation; D002712:Chlorides / Q000494:pharmacology; D003956:Dialysis; D004563:Electrochemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004885:Erwinia / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D050356:Lipid Metabolism; D008055:Lipids / Q000302:isolation & purification; D008070:Lipopolysaccharides / Q000302:isolation & purification / Q000378:metabolism; D046911:Macromolecular Substances; D009994:Osmolar Concentration; D011112:Polymyxin B / Q000494:pharmacology; D011133:Polysaccharide-Lyases / Q000302:isolation & purification*
D002712:Chlorides; D008055:Lipids; D008070:Lipopolysaccharides; D046911:Macromolecular Substances; D011133:Polysaccharide-Lyases; C023911:pectate lyase; D011112:Polymyxin B