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4-Methyl-7-thioumbelliferyl-beta-D-cellobioside: a fluorescent, nonhydrolyzable substrate analogue for cellulases.
- Source :
-
Biochemistry [Biochemistry] 2002 Apr 02; Vol. 41 (13), pp. 4447-52. - Publication Year :
- 2002
-
Abstract
- The kinetics of cellulose binding and hydrolysis by cellulases is not well understood except at steady-state conditions. For use in studies of cellulase pre-steady-state and steady-state kinetics, we have prepared 4-methyl-7-thioumbelliferyl-beta-D-cellobioside (MUS-CB), a ground-state nonhydrolyzable analogue of the fluorescent cellulase substrate 4-methylumbelliferyl-beta-D-cellobioside (MU-CB). MUS-CB is not hydrolyzed by the catalytic domain of cellulase E1 from Acidothermus cellulolyticus under conditions where this enzyme rapidly degrades MU-CB. Thermodynamic parameters describing the steady-state binding of MUS-CB to Thermobifida fusca cellulase Cel6A are similar to those for MU-CB, indicating that MUS-CB can be used in place of MU-CB to study binding events in the Cel6A active-site cleft. In the pre-steady-state, MUS-CB binds to Cel6A by a simple, one-step bimolecular association reaction. It is anticipated that similar thio-containing 4-methylumbelliferyl compounds will have applications in studies of other enzyme systems.
- Subjects :
- Binding Sites
Catalytic Domain
Cellobiose analogs & derivatives
Cellobiose chemistry
Dose-Response Relationship, Drug
Glycosides chemistry
Hydrolases chemistry
Kinetics
Magnetic Resonance Spectroscopy
Models, Chemical
Protein Binding
Thermodynamics
Umbelliferones chemistry
Cellobiose pharmacology
Cellulase chemistry
Fluorescent Dyes pharmacology
Glucosides chemistry
Umbelliferones pharmacology
Subjects
Details
- Language :
- English
- ISSN :
- 0006-2960
- Volume :
- 41
- Issue :
- 13
- Database :
- MEDLINE
- Journal :
- Biochemistry
- Publication Type :
- Academic Journal
- Accession number :
- 11914092
- Full Text :
- https://doi.org/10.1021/bi015854q