Remarkable activation of enzymes in nonaqueous media by denaturing organic cosolvents

✍ Scribed by Örn Almarsson; Alexander M. Klibanov

• Publisher: John Wiley and Sons
• Year: 2000
• Tongue: English
• Weight: 639 KB
• Volume: 49
• Category: Article
• ISSN: 0006-3592
• DOI: 10.1002/(sici)1097-0290(19960105)49:1<87::aid-bit11>3.0.co;2-8

✦ Synopsis

The rates of transesterification reactions catalyzed by the protease subtilisin Carlsberg suspended in various anhydrous solvents at 30°C can be increased more than 100fold by the addition of denaturing organic cosolvents (dimethyl sulfoxide or formamidef; in water, the same cosolvents exert no enzyme activation. A t 4"C, the activation effect on the lyophilized protease is even higher, reaching 1000-fold. Marked enhancement of enzymatic activity in anhydrous solvents by formamide is also observed for two other enzymes, a-chymotrypsin and Rhizomucor miehei lipase, and is manifested in two transesterification reactions. In addition to lyophilized subtilisin, crosslinked crystals of subtilisin are also amenable t o the dramatic activation by the denaturing cosolvents. In contrast, subtilisin solubilized in anhydrous media by covalent modification with poly(ethy1ene glycol) exhibits only modest activation. These observations are rationalized in terms of a mechanistic hypothesis based on an enhanced protein flexibility in anhydrous milieu brought about by the denaturing organic cosolvents. The latter exert their lubricating effect largely at the interfaces between enzyme molecules in a solid preparation, thus easing the flexibility constraints imposed by proteinprotein contacts. 0 1996 John Wiley & Sons, Inc. Key words: protease lipase activation anhydrous