Isotherms for adsorption of cellobiohydrolase I and II from Trichoderma reesei on microcrystalline cellulose.
Adsorption to microcrystalline cellulose (Avicel) of pure cellobiohydrolase I and II (CBH I and CBH II) from Trichoderma reesei has been studied. Adsorption isotherms of the enzymes were measured at 4 degrees C using CBH I and CBH II alone and in reconstituted equimolar mixtures. Several models (Langmuir, Freundlich, Temkin, Jovanovic) were tested to describe the experimental adsorption isotherms. The isotherms did not follow the basic (one site) Langmuir equation that has often been used to describe adsorption isotherms of cellulases; correlation coefficients (R2) were only 0.926 and 0.947, for CBH I and II, respectively. The experimental isotherms were best described by a model of Langmuir type with two adsorption sites and by a combined Langmuir-Freundlich model (analogous to the Hill equation); using these models the correlation coefficients were in most cases higher than 0.995. Apparent binding parameters derived from the two sites Langmuir model indicated stronger binding of CBH II compared to CBH I; the distribution coefficients were 20.7 and 3.7 L/g for the two enzymes, respectively. The binding capacity, on the other hand, was higher for CBH I, 1.0 mumol (67 mg) per gram Avicel, compared to 0.57 mumol/g (30 mg/g) for CBH II. The isotherms when analyzed with the combined Langmuir-Freundlich model indicated presence of unequal binding sites on cellulose and/or negative cooperatively in the binding of the enzyme molecules.
Medve
J
J
Department of Biochemistry, University of Lund, Sweden.
Ståhlberg
J
J
Tjerneld
F
F
D000327:Adsorption; D001665:Binding Sites; D002480:Cellulase / Q000737:chemistry* / Q000302:isolation & purification; D002482:Cellulose; D043366:Cellulose 1,4-beta-Cellobiosidase; D002851:Chromatography, High Pressure Liquid; D005656:Fungal Proteins / Q000737:chemistry* / Q000302:isolation & purification; D006868:Hydrolysis; D007527:Isoenzymes / Q000737:chemistry* / Q000302:isolation & purification; D008956:Models, Chemical; D013816:Thermodynamics; D014242:Trichoderma / Q000201:enzymology*
D005656:Fungal Proteins; D007527:Isoenzymes; D002482:Cellulose; D002480:Cellulase; D043366:Cellulose 1,4-beta-Cellobiosidase