Activity, stability, and conformation of methoxypoly(ethylene glycol)-subtilisin at different concentrations of water in dioxane

摘要: The transesterification activity, autolysis, thermal stability and conformation of subtilisin Carlsberg, made soluble in dioxane by covalent linking to methoxypoly(ethylene glycol) (PEG), were investigated as a function the concentration water medium. Electrospray mass spectrometry showed that modified enzyme preparation was mixture proteins containing from 2 5 covalently linked PEG chains per molecule. PEG-subtilisin catalyzed between vinyl butyrate 1-hexanol optimum at 0.55 MH2O, while hydrolysis prevailed above MH2O. There decrease overall activity with increasing because autolysis denaturation enzyme. Subtilisin powder celite-immobilized more stable less susceptible than PEG-modified Circular dichroism intrinsic protein-fluorescence studies did not change concentrations 0 9 M. Km,app value for highly influenced water, which behaved competitive inhibitor reaction an affinity similar alcohol. acylating agent significantly water. Lyoprotectants such sorbitol free influence but notably increased subtilisin. addition 1.7–5.5 M however, rendered preparations no additives active those lyoprotectants. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng54: 50–57, 1997.