# Atomic resolution analysis of timekeeping by a protein-based circadian clock

> **NIH NIH R01** · UNIVERSITY OF CALIFORNIA SANTA CRUZ · 2020 · $367,742

## Abstract

Project Summary
The ability to encode time at the molecular level is universally used to synchronize biological processes into
~24-hour rhythms that coincide with the solar day for health and homeostasis. How do we measure time at the
molecular level and use it as the basis for biological regulation? The goal of the current project is to use the
simplest model system for circadian cycling to explore the structural basis for generating a molecular clock that
keeps 24-hour time. Cyanobacteria have a biochemically tractable clock composed of three Kai (cycle)
proteins that keep ~24-hour time in vitro, requiring only the addition of ATP as an energy source. KaiC is the
central pacemaker of this clock, with two tandem ATPase domains (C1 and C2) that each assemble into
hexameric rings connected by a flexible linker. Information about the time of day is transmitted between the
rings to influence interactions with KaiA and KaiB, as well as with proteins that transmit information about the
environment to the clock and control biological timing in vivo. Despite its relatively simple composition, the lack
of structures for intermediate states formed by Kai protein complexes has limited our understanding of this
molecular clock. The proposed research will provide structural, biochemical, and in vivo functional data in
support of a new model for Kai protein interactions, leading to a major shift in our understanding of the
cyanobacterial circadian oscillator. We are pursuing a hypothesis that competition for the KaiC C1 domain, a
central hub for clock protein interactions, is essential for circadian timekeeping. Our approach is built on the
discovery that KaiB undergoes a metamorphic transition to a new protein fold that is needed to regulate
assembly of clock protein complexes with KaiC. Using a version of KaiB locked into its rare, active
conformation, we present three new structures of clock protein complexes that demonstrate the vastly
underappreciated role that the KaiC C1 domain plays in generating the molecular clock. In Aim 1, we will
determine how the hexameric KaiC ATPase controls assembly of Kai protein complexes. In Aim 2, we will
identify the structural basis for communication between the two ATPase rings of KaiC. In Aim 3, we will
determine how competitive interactions at the KaiC C1 domain control clock signaling throughout the day and
night. The proposed studies will demonstrate the structural basis by which KaiC integrates interactions with
clock proteins to keep 24-hour time and transform our understanding of the cyanobacterial circadian clock.

## Key facts

- **NIH application ID:** 9856464
- **Project number:** 5R01GM121507-04
- **Recipient organization:** UNIVERSITY OF CALIFORNIA SANTA CRUZ
- **Principal Investigator:** Carrie L Partch
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $367,742
- **Award type:** 5
- **Project period:** 2017-02-01 → 2023-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9856464

## Citation

> US National Institutes of Health, RePORTER application 9856464, Atomic resolution analysis of timekeeping by a protein-based circadian clock (5R01GM121507-04). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9856464. Licensed CC0.

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