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Coupling ATP utilization to protein remodeling by ClpB, a hexameric AAA+ protein
Joel R Hoskins1, Shannon M Doyle, Sue Wickner
1Laboratory of Molecular Biology, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Summary
ClpB protein remodeling activity depends on ATP. Without the DnaK system, ClpB uses a probabilistic mechanism, but with DnaK, it follows a sequential mechanism.
Area of Science:
- Molecular Biology
- Protein Biochemistry
- Cellular Stress Response
Background:
- ClpB and Hsp104 are AAA+ ATPases crucial for thermotolerance.
- These proteins remodel proteins but require the DnaK/Hsp70 system for disaggregation.
- The precise mechanism of ClpB's ATP utilization in protein remodeling is not fully understood.
Purpose of the Study:
- To investigate the mechanism of ClpB's ATP coupling to protein remodeling.
- To determine how the DnaK system influences ClpB's mechanism.
- To elucidate the adaptability of ClpB's ATP utilization.
Main Methods:
- Constructed ClpB heterohexamers with wild-type and ATP hydrolysis-deficient protomers.
- Assessed protein remodeling activity with and without the DnaK system.
- Analyzed the impact of active/inactive protomer ratios and ATP binding site location.
Main Results:
- ClpB heterohexamers lacking DnaK gained remodeling activity, suggesting a probabilistic mechanism.
- Optimal activity without DnaK required a 3:3 ratio of active to inactive protomers.
- In the presence of DnaK, even one inactive ClpB subunit blocked activity, indicating a sequential mechanism.
Conclusions:
- ClpB's ATP utilization mechanism is adaptable.
- The mechanism shifts from probabilistic (without DnaK) to sequential (with DnaK).
- Substrate and chaperone presence dictate ClpB's mechanistic strategy.
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