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Related Experiment Video

Updated: Jan 19, 2026

Measurement of Tissue Non-Heme Iron Content using a Bathophenanthroline-Based Colorimetric Assay
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Heme binding to human CLOCK affects interactions with the E-box.

Samuel L Freeman1, Hanna Kwon1, Nicola Portolano2,3

  • 1School of Chemistry, University of Bristol, BS8 1TS Bristol, United Kingdom.

Proceedings of the National Academy of Sciences of the United States of America
|September 19, 2019
PubMed
Summary
This summary is machine-generated.

Heme binding to the human CLOCK protein disrupts its DNA binding, revealing a novel mechanism for heme-dependent circadian rhythm regulation.

Keywords:
CLOCKcircadianheme

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Chronobiology

Background:

  • The circadian clock regulates biological rhythms.
  • CLOCK and BMAL1 are key mammalian circadian clock proteins.
  • Heme's role in circadian control is not well understood.

Purpose of the Study:

  • To investigate the interaction between heme and human CLOCK (hCLOCK).
  • To elucidate the structural and functional consequences of heme binding to hCLOCK.

Main Methods:

  • X-ray crystallography to determine the structure of hCLOCK's PAS-A domain.
  • UV-visible and electron paramagnetic resonance spectroscopies to study heme binding.
  • Site-directed mutagenesis to identify heme-ligating residues.
  • DNA binding assays to assess hCLOCK-E-box interactions.

Main Results:

  • A crystal structure of the hCLOCK PAS-A domain was determined.
  • Heme binds to the PAS-A domain, with His144 identified as a key ligand.
  • Heme binding alters hCLOCK conformation and disrupts its binding to the E-box DNA target.
  • Evidence suggests a flexible heme pocket with potential His/Cys coordination.

Conclusions:

  • Heme directly interacts with hCLOCK, impacting its function.
  • Heme binding to hCLOCK provides a mechanism for heme-dependent transcriptional regulation of circadian rhythms.
  • The conformational flexibility of hCLOCK is linked to heme ligation and DNA binding efficacy.