A novel and simple method for the purification of extracellular levansucrase from Zymomonas mobilis.
A new and simple method for the purification of extracellular levansucrase from Zymomonas mobilis from highly viscous fermentation broth was developed. After incubation of the fermentation broth with a fructose-polymer cleaving enzyme preparation (Fructozyme, Novozymes, DK) for 48 h, levansucrase precipitated as aggregates and was redissolved in a 3 M urea solution. By ongoing size-exclusion chromatography on Sephacryl S-300 the final levansucrase preparation was purified 100-fold and exhibited a specific activity of 25-35 U/mg(protein). The levansucrase was stable in 3 M urea solution for at least four months without inactivation. To maximize the enzyme yield the dynamic changes of extracellular levansucrase activity during fermentation were investigated. The highest levansucrase activity was observed during the logarithmic phase of growth (15-19 h of fermentation).
Institute of Microbiology and Biotechnology, University of Latvia, Kronvalda Boulevard 4, 1586 Riga, Latvia. avigants@lanet.LV
Armands
A
Vigants
Stefan Peter
SP
Marx
Raimonds
R
Linde
Solvita
S
Ore
Martins
M
Bekers
Ilmara
I
Vina
Hans-Georg
HG
Hicke
D002498:Centrifugation; D011232:Chemical Precipitation; D002850:Chromatography, Gel; D003470:Culture Media / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005285:Fermentation; D005630:Fructans / Q000378:metabolism; D006602:Hexosyltransferases / Q000302:isolation & purification*; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D012995:Solubility; D013395:Sucrose / Q000378:metabolism; D014508:Urea; D014783:Viscosity; D017355:Zymomonas / Q000201:enzymology*
D003470:Culture Media; D005630:Fructans; D013395:Sucrose; D014508:Urea; C072599:levan; D006602:Hexosyltransferases; C021513:levansucrase