[Synthesis and properties of immobilized enzymes. X. Covalent binding of polygalacturonase to insoluble carriers].
The pectinolytic enzymes are of practical interest for the clarification of fruit juice. In the present paper the covalent coupling of polygalacturonase (PG; E. C. 3.2.1.15) is reported. A commercially available enzyme (Rohament P; 5 U/mg) and purified Endo-PG (200 U/mg) are immobilized to the following carriers: BrCN-activated Sepharose, carbodiimide-activated CH-Sepharose, dialdehyde Sepharose, dialdehyde Sephadex, dialdehyde cellulose, CMC-azide, carbodiimide-activated CMC, macroporous glass (isothiocyanate and carbodiimide coupling) and glass beads. The implications of pore diameter (Sephadex- and Sepharose derivatives), of purity of the PG, of protein content of the PG-carrier-complexes as well as the presence of substrate during the coupling reaction, are discused in relation to the relative and specific activity of the bound protein and to the efficiency of the coupling reaction. From the carriers under study derivatives of Sepharose yield the best result (relative activity max. 88%, specific activity max. 5400 U/mg). The immobilization to isothiocyanate glass yields good results, too (relative activity 20%, specific activity 500 U/g). The mechanical instability of the PG-dialdehye Sephadex-complexes and the low relative activity of the bound enzyme are unsatisfactory. Due to their low affinity to PG, the derivatives of cellulose are also inappropriate for covalent coupling of this enzyme. All PG-carrier-complexes are largely stable both during storage at 4 degrees C and repeated activity assays.
D004800:Enzymes, Immobilized / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000378:metabolism*; D011096:Polygalacturonase / Q000378:metabolism*; D011485:Protein Binding; D012685:Sepharose; D013329:Structure-Activity Relationship
D004800:Enzymes, Immobilized; D012685:Sepharose; D006026:Glycoside Hydrolases; D011096:Polygalacturonase
Synthese und Eigenschaften trägerfixierter Enzyme. X. Kovalente Bindung von Polygalakturonase an wasserunlösliche Träger.