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Ordered self-assembly of polypeptide fragments to form nativelike dimeric trp repressor
1Chemistry Department, Princeton University, NJ 08544.
Science (New York, N.Y.)
|January 31, 1992
Summary
Researchers created protein fragments that reassemble into functional Escherichia coli trp repressor. This method reveals key folding intermediates, supporting the link between protein structure stability and folding pathways.
Area of Science:
- Molecular Biology
- Biochemistry
- Structural Biology
Background:
- Understanding protein folding mechanisms is crucial for deciphering protein function and dysfunction.
- The Escherichia coli trp repressor is a well-studied model system for protein folding.
- Spectroscopic methods provide kinetic data but limited structural detail on folding intermediates.
Purpose of the Study:
- To investigate the folding pathway of the Escherichia coli trp repressor using a novel fragment reassembly approach.
- To characterize the structure of transient folding intermediates.
- To correlate structural data with kinetic folding rates.
Main Methods:
- Production of subdomain-size proteolytic fragments of the trp repressor.
- Assembly of fragments to regenerate native repressor dimers.
- Characterization of secondary and tertiary structures of fragments and reassembled intermediates using spectroscopic techniques.
- Kinetic analysis of folding rates via spectroscopic measurements.
Main Results:
- Defined order assembly of proteolytic fragments to regenerate native trp repressor dimers.
- Characterization of secondary and tertiary structures of isolated and recombined fragments.
- Correlation of intermediate structures with the kinetic folding pathway.
- Nativelike structure observed in folding intermediates.
Conclusions:
- Protein folding pathways reflect the stability of secondary structural units and native-state assemblies.
- The proteolytic fragment reassembly method provides structural detail to folding mechanisms.
- This approach offers a general strategy for studying protein folding pathways.
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