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Rhodanese folding is controlled by the partitioning of its folding intermediates
B M Gorovits1, W A McGee, P M Horowitz
1Department of Biochemistry, University of Texas Health Science Center at San Antonio 78284, USA.
Biochimica Et Biophysica Acta
|March 21, 1998
Summary
Investigating bovine rhodanese refolding revealed that glycerol stabilizes folding intermediates, preventing aggregation and increasing refolding yield. This suggests associative processes and domain dissociation are key to enzyme renaturation.
Area of Science:
- Biochemistry
- Protein Folding
- Enzymology
Background:
- Rhodanese is a model enzyme for protein folding studies.
- Unassisted refolding of rhodanese is often inefficient.
- Metastable intermediates form during rhodanese unfolding.
Purpose of the Study:
- To investigate how partitioning of folding intermediates affects bovine rhodanese refolding efficiency.
- To understand the role of glycerol in stabilizing intermediates and promoting refolding.
- To explore the nature of aggregation-prone intermediates.
Main Methods:
- Chemical unfolding and refolding kinetics analysis.
- High glycerol concentration studies to assess intermediate stability.
- Investigation of interactions with the chaperonin GroEL.
Main Results:
- Glycerol significantly increases the stability of folding intermediates and native rhodanese.
- Protein folding kinetics suggest associative processes during renaturation.
- Rhodanese collapses to an intermediate state that partitions into refoldable and aggregation-prone states.
- Stabilization of the aggregation-prone intermediate by glycerol enhances refolding yield.
- Extensively unfolded intermediates bind more strongly to GroEL.
Conclusions:
- Glycerol enhances rhodanese refolding by stabilizing intermediates and preventing aggregation.
- Associative mechanisms and potential domain dissociation are involved in rhodanese folding.
- The chaperonin GroEL interacts differentially with unfolding intermediates based on their extent of unfolding.