Directed Evolution of Substrate-Optimized GroEL/S Chaperonins

GroEL/S chaperonin ring complexes fold many unrelated proteins. To understand the basis and extent of the chaperonin substrate spectrum, we used rounds of selection and DNA shuffling to obtain GroEL/S variants that dramatically enhanced folding of a single substrate-green fluorescent protein (GFP)....

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Bibliographic Details
Published in:Cell 2002-12, Vol.111 (7), p.1027-1039
Main Authors: Wang, Jue D., Herman, Christophe, Tipton, Kimberly A., Gross, Carol A., Weissman, Jonathan S.
Format: Article
Language:English
Subjects:
Adenosine Triphosphatases - genetics
Adenosine Triphosphatases - metabolism
Chaperonin 10 - genetics
Chaperonin 10 - metabolism
Chaperonin 60 - genetics
Chaperonin 60 - metabolism
Directed Molecular Evolution
Escherichia coli
Eukaryotic Cells - metabolism
Evolution, Molecular
Gene Expression Regulation, Bacterial - genetics
Green Fluorescent Proteins
Luminescent Proteins - genetics
Luminescent Proteins - metabolism
Mutation - genetics
Prokaryotic Cells - metabolism
Protein Folding
Protein Isoforms - genetics
Protein Isoforms - metabolism
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Summary:GroEL/S chaperonin ring complexes fold many unrelated proteins. To understand the basis and extent of the chaperonin substrate spectrum, we used rounds of selection and DNA shuffling to obtain GroEL/S variants that dramatically enhanced folding of a single substrate-green fluorescent protein (GFP). Changes in the substrate-optimized chaperonins increase the polarity of the folding cavity and alter the ATPase cycle. These findings reveal a surprising plasticity of GroEL/S, which can be exploited to aid folding of recombinant proteins. Our studies also reveal a conflict between specialization and generalization of chaperonins as increased GFP folding comes at the expense of the ability of GroEL/S to fold its natural substrates. This conflict and the nature of the ring structure may help explain the evolution of cellular chaperone systems.
ISSN:0092-8674
1097-4172
DOI:10.1016/S0092-8674(02)01198-4