Facial reconstruction

Search LJMU Research Online

Browse Repository | Browse E-Theses

Heme binding to human CLOCK affects interactions with the E-box.

Freeman, SL, Kwon, H, Portolano, N, Parkin, G, Venkatraman Girija, U, Basran, J, Fielding, AJ, Fairall, L, Svistunenko, DA, Moody, PCE, Schwabe, JWR, Kyriacou, CP and Raven, EL (2019) Heme binding to human CLOCK affects interactions with the E-box. Proceedings of the National Academy of Sciences, 40. pp. 19911-19916. ISSN 0027-8424

[img] Text
Raven et al for green open access.pdf - Accepted Version
Restricted to Repository staff only until 16 March 2020.

Download (15MB)

Abstract

The circadian clock is an endogenous time-keeping system that is ubiquitous in animals and plants as well as some bacteria. In mammals, the clock regulates the sleep-wake cycle via 2 basic helix-loop-helix PER-ARNT-SIM (bHLH-PAS) domain proteins-CLOCK and BMAL1. There is emerging evidence to suggest that heme affects circadian control, through binding of heme to various circadian proteins, but the mechanisms of regulation are largely unknown. In this work we examine the interaction of heme with human CLOCK (hCLOCK). We present a crystal structure for the PAS-A domain of hCLOCK, and we examine heme binding to the PAS-A and PAS-B domains. UV-visible and electron paramagnetic resonance spectroscopies are consistent with a bis-histidine ligated heme species in solution in the oxidized (ferric) PAS-A protein, and by mutagenesis we identify His144 as a ligand to the heme. There is evidence for flexibility in the heme pocket, which may give rise to an additional Cys axial ligand at 20K (His/Cys coordination). Using DNA binding assays, we demonstrate that heme disrupts binding of CLOCK to its E-box DNA target. Evidence is presented for a conformationally mobile protein framework, which is linked to changes in heme ligation and which has the capacity to affect binding to the E-box. Within the hCLOCK structural framework, this would provide a mechanism for heme-dependent transcriptional regulation.

Item Type: Article
Uncontrolled Keywords: CLOCK; circadian; heme
Subjects: Q Science > QH Natural history > QH301 Biology
Q Science > QH Natural history > QH426 Genetics
Divisions: Pharmacy & Biomolecular Sciences
Publisher: National Academy of Sciences
Related URLs:
Date Deposited: 02 Oct 2019 08:35
Last Modified: 02 Oct 2019 08:35
DOI or Identification number: 10.1073/pnas.1905216116
URI: http://researchonline.ljmu.ac.uk/id/eprint/11419

Actions (login required)

View Item View Item