Effect of directional pulling on mechanical protein degradation by ATP-dependent proteolytic machines
AAA+ proteases and remodeling machines couple hydrolysis of ATP to mechanical unfolding and translocation of proteins following recognition of sequence tags called degrons. Here, we use single-molecule optical trapping to determine the mechanochemistry of two AAA+ proteases, Escherichia coli ClpXP a...
| Main Authors: | , , , , |
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| Other Authors: | |
| Format: | Article |
| Published: |
National Academy of Sciences (U.S.)
2018
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| Online Access: | http://hdl.handle.net/1721.1/114914 https://orcid.org/0000-0001-9751-9535 https://orcid.org/0000-0002-2246-2674 https://orcid.org/0000-0002-1719-5399 |
| Summary: | AAA+ proteases and remodeling machines couple hydrolysis of ATP to mechanical unfolding and translocation of proteins following recognition of sequence tags called degrons. Here, we use single-molecule optical trapping to determine the mechanochemistry of two AAA+ proteases, Escherichia coli ClpXP and ClpAP, as they unfold and translocate substrates containing multiple copies of the titin[superscript I27] domain during degradation initiated from the N terminus. Previous studies characterized degradation of related substrates with C-terminal degrons. We find that ClpXP and ClpAP unfold the wild-type titin I27 domain and a destabilized variant far more rapidly when pulling from the N terminus, whereas translocation speed is reduced only modestly in the N-to-C direction. These measurements establish the role of directionality in mechanical protein degradation, show that degron placement can change whether unfolding or translocation is rate limiting, and establish that one or a few power strokes are sufficient to unfold some protein domains. Keywords:protein degradation; AAA+ proteases; directional unfolding; AAA+ motors |
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