Immobilized enzymes in reverse micelles: Studies with gel-entrapped trypsin and α-chymotrypsin in AOT reverse micelles

✍ Scribed by Nitin W. Fadnavis; Pier Luigi Luisi

• Publisher: John Wiley and Sons
• Year: 1989
• Tongue: English
• Weight: 564 KB
• Volume: 33
• Category: Article
• ISSN: 0006-3592
• DOI: 10.1002/bit.260331008

✦ Synopsis

Trypsin and a-chymotrypsin were immobilized by gel entrapment in polyacrylamide cross-linked with N , Nmethylenebisacrylamide. The immobilized enzymes are catalytically efficient in suspensions of reverse micelles formed i n isooctane by bis(2-ethylhexyl) sodium sulfosuccinate) (AOT) and water. Both entrapped enzymes are stable in reverse micellar suspension at room temperature and pH 8.2 for 3 days and lose 30-40% activity after 1 week. The enzymes obey Michaelis-Menten kinetics in the investigated concentration range with K,,, values higher than those in solution. Activity of the enzymes is independent of the water content of the micellar solution. No shift in pH optimum was observed for immobilized trypsin activity toward Na-benzoyl-Larginine ethyl ester. The utility of the procedure, which combines the advantage of enzyme immobilization and enzymology i n reverse micelles, is illustrated by an example of peptide synthesis. In particular, peptide synthesis (e.g., Z-Ala-Phe-Leu-NH,) using water-insoluble substrate has been performed with gelentrapped a-chymotrypsin in reverse micellar suspension with the advantage of efficient enzyme recycling.