# Structure and function of ClpXP

> **NIH NIH R35** · MASSACHUSETTS INSTITUTE OF TECHNOLOGY · 2021 · $385,688

## Abstract

Project Summary
AAA+ proteases remove toxic proteins and regulate many other important cellular processes
that promote health and prevent disease. At the same time, protein degradation must be
carefully regulated. AAA+ proteases assemble into multi-subunit structures with an internal
proteolytic chamber, accessible through narrow channels that exclude natively folded proteins.
This mechanism protects most proteins from unintended degradation and requires specific
substrates to be recognized, unfolded, and then translocated into the degradation chamber. In
the AAA+ ClpXP protease, for example, a ring hexamer of ClpX uses the energy of ATP
hydrolysis to unfold specific target proteins and translocate them into ClpP for degradation.
ClpXP is one of the best-characterized AAA+ proteases and is a paradigm for other ATP-
dependent proteases and AAA+ remodeling machines. These ATP-fueled enzymes perform a
wide variety of mechanical remodeling, transport, and regulatory tasks in the cell. In mammals,
loss of mitochondrial ClpP results in infertility, hearing loss, and growth defects, whereas
mitochondrial ClpX plays an important role in heme biosynthesis. Bacterial ClpXP can promote
pathogenesis and is a validated antibiotic target in M. tuberculosis. Substantial progress has
been made in understanding the general biochemical and structural features of E. coli ClpXP
and other AAA+ enzymes but important and fundamental questions concerning the molecular
mechanisms of these machines remain. For example, it is not known how ClpX identifies many
classes of N-terminal and C-terminal degrons, whether ClpX rotates with respect to ClpP
during normal function, whether proofreading helps ensure degradation specificity, how
multiple substrate chains can be simultaneously translocated through the ClpX channel, what
the detailed and interacts with polypeptide substrates during mechanical unfolding, whether
ATP hydrolysis can occur at multiple positions in the spiral ClpX ring or only at one or a few
special positions, and how the detailed ATPase cycle is coupled to mechanical work. The
experiments described in this proposal will address these questions and provide a conceptual
framework applicable to studies of the entire superfamily of AAA+ machines.

## Key facts

- **NIH application ID:** 10198307
- **Project number:** 1R35GM141517-01
- **Recipient organization:** MASSACHUSETTS INSTITUTE OF TECHNOLOGY
- **Principal Investigator:** Robert T Sauer
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $385,688
- **Award type:** 1
- **Project period:** 2021-04-01 → 2026-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10198307

## Citation

> US National Institutes of Health, RePORTER application 10198307, Structure and function of ClpXP (1R35GM141517-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10198307. Licensed CC0.

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