Purification of glucosyltransferase from cell-lysate of Streptococcus mutans by counter-current chromatography using aqueous polymer two-phase system

Counter-current chromatography (CCC) using a cross-axis coil planet centrifuge (X-axis CPC) was applied to the purification of glucosyltransferase (GTF) from a cell-lysate of cariogenic bacteria. The purification was performed using an aqueous polymer two-phase system composed of 4.4% (w/w) polyethy...

Full description

Saved in:
Bibliographic Details
Published in:Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Analytical technologies in the biomedical and life sciences, 2004-06, Vol.805 (1), p.155-160
Main Authors: Yanagida, Akio, Isozaki, Mitsuhiro, Shibusawa, Yoichi, Shindo, Heisaburo, Ito, Yoichiro
Format: Article
Language:English
Subjects:
Analysis
Analytical, structural and metabolic biochemistry
Biological and medical sciences
Chromatography, Liquid - methods
Countercurrent Distribution - methods
Electrophoresis, Polyacrylamide Gel
Fundamental and applied biological sciences. Psychology
General pharmacology
Glucosyltransferase
Glucosyltransferases - isolation & purification
Medical sciences
Pharmacology. Drug treatments
Spectrometry, Mass, Electrospray Ionization
Streptococcus mutans
Streptococcus mutans - enzymology
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Counter-current chromatography (CCC) using a cross-axis coil planet centrifuge (X-axis CPC) was applied to the purification of glucosyltransferase (GTF) from a cell-lysate of cariogenic bacteria. The purification was performed using an aqueous polymer two-phase system composed of 4.4% (w/w) polyethylene glycol (PEG) 8000–6% (w/w) dextran T500 containing 10 mM phosphate buffer at pH 9.2 by eluting the upper phase (UP) at 1.0 ml/min. The bacterial GTF in the cell-lysate of Streptococcus mutans was selectively retained in the dextran-rich lower stationary phase. The column contents were diluted and subjected to hydroxyapatite (HA) chromatography to remove the polymers from the GTF. Fractions eluted with 500 mM potassium phosphate buffer were analyzed by GTF enzymatic activity as well as sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS–PAGE). The GTF purity in the final product was increased about 87 times as that in the cell-lysate with a good recovery rate of about 79% through this purification process.
ISSN:1570-0232
1873-376X
DOI:10.1016/j.jchromb.2004.02.039