An empirical energy landscape reveals mechanism of proteasome in polypeptide translocation
Curation statements for this article:-
Curated by eLife
Evaluation Summary:
AAA+ ATPases consume chemical energy in form of ATP to catalyze essential cellular reactions. Here, computational and biochemical approaches are used to model how the six subunits of the AAA+ ATPase Rpt1-6 coordinate their enzymatic activity with each other to exert unidirectional pulling forces on target polypeptide chains that promote protein unfolding. Although the technical aspects of the work are sometimes difficult to follow, the findings indicate that the order in which ATPase active sites fire is generally sequential, much like a rotary engine. The system can tolerate "misfires" - instances in which a subunit fails to hydrolyze ATP - by skipping the problematic subunit. The work should appeal to the broad AAA+ community and researchers trying to understand the biophysical principles by which complex biological machines operate.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #3 agreed to share their name with the authors.)
This article has been Reviewed by the following groups
Listed in
- Evaluated articles (eLife)
- Structural Biology and Molecular Biophysics (eLife)
Abstract
The ring-like ATPase complexes in the AAA+ family perform diverse cellular functions that require coordination between the conformational transitions of their individual ATPase subunits (Erzberger and Berger, 2006; Puchades et al., 2020). How the energy from ATP hydrolysis is captured to perform mechanical work by these coordinated movements is unknown. In this study, we developed a novel approach for delineating the nucleotide-dependent free-energy landscape (FEL) of the proteasome’s heterohexameric ATPase complex based on complementary structural and kinetic measurements. We used the FEL to simulate the dynamics of the proteasome and quantitatively evaluated the predicted structural and kinetic properties. The FEL model predictions are consistent with a wide range of experimental observations in this and previous studies and suggested novel mechanistic features of the proteasomal ATPases. We find that the cooperative movements of the ATPase subunits result from the design of the ATPase hexamer entailing a unique free-energy minimum for each nucleotide-binding status. ATP hydrolysis dictates the direction of substrate translocation by triggering an energy-dissipating conformational transition of the ATPase complex.
Article activity feed
-
-
Evaluation Summary:
AAA+ ATPases consume chemical energy in form of ATP to catalyze essential cellular reactions. Here, computational and biochemical approaches are used to model how the six subunits of the AAA+ ATPase Rpt1-6 coordinate their enzymatic activity with each other to exert unidirectional pulling forces on target polypeptide chains that promote protein unfolding. Although the technical aspects of the work are sometimes difficult to follow, the findings indicate that the order in which ATPase active sites fire is generally sequential, much like a rotary engine. The system can tolerate "misfires" - instances in which a subunit fails to hydrolyze ATP - by skipping the problematic subunit. The work should appeal to the broad AAA+ community and researchers trying to understand the biophysical principles by which complex …
Evaluation Summary:
AAA+ ATPases consume chemical energy in form of ATP to catalyze essential cellular reactions. Here, computational and biochemical approaches are used to model how the six subunits of the AAA+ ATPase Rpt1-6 coordinate their enzymatic activity with each other to exert unidirectional pulling forces on target polypeptide chains that promote protein unfolding. Although the technical aspects of the work are sometimes difficult to follow, the findings indicate that the order in which ATPase active sites fire is generally sequential, much like a rotary engine. The system can tolerate "misfires" - instances in which a subunit fails to hydrolyze ATP - by skipping the problematic subunit. The work should appeal to the broad AAA+ community and researchers trying to understand the biophysical principles by which complex biological machines operate.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #3 agreed to share their name with the authors.)
-
Reviewer #1 (Public Review):
Ring-shaped proteins known as AAA+ ATPases consume chemical energy in form of ATP to catalyze essential cellular reactions ranging from the copying and repair of DNA to signal transmission at neuronal synapses. In the present study, computational and biochemical approaches are used to model how the six subunits of a particular AAA+ ATPase known as Rpt1-6 coordinate their enzymatic activity with each other to exert unidirectional pulling forces on target polypeptide chains that promote protein unfolding. Although the technical aspects of the work can be difficult to follow at times, the conclusions seem largely supported by the observations. The findings indicate that the order in which ATPase active sites fire around the is generally sequential, much like a rotary engine, but that the system can tolerate …
Reviewer #1 (Public Review):
Ring-shaped proteins known as AAA+ ATPases consume chemical energy in form of ATP to catalyze essential cellular reactions ranging from the copying and repair of DNA to signal transmission at neuronal synapses. In the present study, computational and biochemical approaches are used to model how the six subunits of a particular AAA+ ATPase known as Rpt1-6 coordinate their enzymatic activity with each other to exert unidirectional pulling forces on target polypeptide chains that promote protein unfolding. Although the technical aspects of the work can be difficult to follow at times, the conclusions seem largely supported by the observations. The findings indicate that the order in which ATPase active sites fire around the is generally sequential, much like a rotary engine, but that the system can tolerate "misfires" - instances in which a subunit fails to hydrolyze ATP - by skipping the problematic subunit. The work should appeal to the broad AAA+ community and researchers trying to understand the biophysical principles by which complex biological machines operate.
-
Reviewer #2 (Public Review):
I found the manuscript difficult to follow and assess in part because of the vast scope of the experimental approach. At the same time, the conclusions could be worked out more clearly and put into the context of the current understanding of proteasome action.
-
Reviewer #3 (Public Review):
This paper presents a simple mathematical model that explains how conformational dynamics of the 26S proteosome enable it to perform mechanical work. The model/approach also have broad implications for other members of the AAA+ family of motors, adding to the significance of the results. Despite a number of simplifying assumptions (e.g. that all 6 subunits have identical behavior), the model does remarkably well at reproducing a variety of experimental data. In particular, the model recapitulates the dominant states/transitions seen experimentally without building in serial transitions. Other transitions are possible, they just have lower probability. Importantly, this heterogeneity of possible pathways allows the motors to continue functioning even when deleterious mutations are introduced, in agreement …
Reviewer #3 (Public Review):
This paper presents a simple mathematical model that explains how conformational dynamics of the 26S proteosome enable it to perform mechanical work. The model/approach also have broad implications for other members of the AAA+ family of motors, adding to the significance of the results. Despite a number of simplifying assumptions (e.g. that all 6 subunits have identical behavior), the model does remarkably well at reproducing a variety of experimental data. In particular, the model recapitulates the dominant states/transitions seen experimentally without building in serial transitions. Other transitions are possible, they just have lower probability. Importantly, this heterogeneity of possible pathways allows the motors to continue functioning even when deleterious mutations are introduced, in agreement with experiments. Deleterious mutations are modeled by removing key interactions in the model, rather than explicitly modeling the physical chemistry of different amino acids. This example highlights how the model can be used to explore the motors conceptually, but is not suited to predicting the impact of specific mutations (or other perturbations). The authors also do a reasonable job of acknowledging the limitations of their model.
-
