Adsorption of Kluyveromyces marxianus pectinase on CM-Sephadex gels.
New results are presented on the equilibria and kinetics of the ion exchange onto CM-Sephadex of polygalacturonase (pectinase) produced by the fermentation of Kluyveromyces marxianus. It is found that the equilibrium behavior follows the form of the Langmuir isotherm; the equilibrium is strongly affected by pH. High partitioning onto the ion-exchange matrix, with good retention of enzyme activity, is achieved in the pH range 3.5-5.0, and this can be qualitatively explained in terms of simple models for protein adsorption by ion exchange. The kinetics of ion exchange is modeled by assuming that the transfer resistances can be lumped into a single coefficient, and the results show that this gives a reasonable description of the adsorption kinetics. Under optimum conditions protein adsorption is enhanced by electrostatic effects and is extremely fast, and it is suggested that in these circumstances external mass transfer resistance is significant. At pH values close to the isoelectric point, electrostatic interactions are weak and intraparticle diffusion is rate-limiting: pore-blocking by adsorbed proteins appears to be important under these conditions. The results also provide the basis for an efficient single-step purification scheme.
Harsa
S
S
Department of Food Science and Technology, University of Reading, UK.
Zaror
C A
CA
Pyle
D L
DL
D000042:Absorption; D002852:Chromatography, Ion Exchange; D003911:Dextrans / Q000737:chemistry*; D005285:Fermentation; D005782:Gels; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007716:Kluyveromyces / Q000201:enzymology* / Q000502:physiology; D009994:Osmolar Concentration; D011096:Polygalacturonase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism
D003911:Dextrans; D005782:Gels; C025614:sephadex; D011096:Polygalacturonase