Structural basis for human PHF2 Jumonji domain interaction with metal ions

• Type of Material: Article
• Authors: John Horton, Emory University; Anup K. Upadhyay, Emory University; Hideharu Hashimoto, Emory University; Xing Zhang, Emory University; Xiaodong Cheng, Emory University
• Language: English
• Date: 2011-02-11
• Publisher: Elsevier
• Publication Version: Author Accepted Manuscript (After Peer Review)
• Copyright Statement: © 2010 Elsevier Ltd. All rights reserved.
• License: Creative Commons Attribution-NonCommerical-NoDerivs 3.0 Unported
• Final Published Version (URL): http://dx.doi.org/10.1016/j.jmb.2010.12.013
• Title of Journal or Parent Work: Journal of Molecular Biology
• ISSN: 0022-2836
• Volume: 406
• Issue: 1
• Start Page: 1
• End Page: 8
• Grant/Funding Information: X.C. is a Georgia Research Alliance Eminent Scholar.; The Department of Biochemistry at the Emory University School of Medicine supported the use of the SER-CAT synchrotron beamline at the Advanced Photon Source of Argonne National Laboratory, local X-ray facility and MALDI-TOF mass spectrometry.; This work was supported by grants GM068680 to X.C. from the US National Institutes of Health.
• Keywords: Epe1; histone lysine demethylation; epigenetics; PHF2; methyl-lysine binding
• Research Categories: Biology, Molecular; Chemistry, Biochemistry

Abstract

PHF2 belongs to a class of α-ketoglutarate-Fe2+–dependent dioxygenases. PHF2 harbors a plant homeodomain (PHD) and a Jumonji domain. PHF2, via its PHD domain, binds Lys4-trimethylated histone 3 (H3K4me3) in sub micromolar affinity, and has been reported to have demethylase activity of mono-methylated lysine 9 of histone 3 (H3K9me1) in vivo. However, we did not detect demethylase activity for PHF2 Jumonji domain (with and without its linked PHD) in the context of histone peptides. We determined the crystal structures of PHF2 Jumonji domain in the absence and presence of additional exogenous metal ions. Only when exposed to high metal concentration (50 mM), Fe2+ or Ni2+ was soaked into the preformed crystals and bound by six ligands in an octahedral coordination. The side-chains of H249, D251, and the two oxygen atoms of N-oxalylglycine (an analog of α-ketoglutarate) provide four coordinations in the equatorial plane, while the hydroxyl oxygen atom of Y321 and one water molecule provide the two axial coordinations as the fifth and sixth ligands, respectively. The metal binding site in PHF2 closely resembles the Fe2+ sites in other Jumonji domains examined, with one important difference, that a tyrosine (Y321 of PHF2) replaces a histidine as the fifth ligand. However, neither Y321H mutation nor high metal concentration renders PHF2 an active demethylase on histone peptides. Both wild type and Y321H mutant bind Ni2+ with approximately equal affinity of 50 µM. We propose there must be other regulatory factors required for the enzymatic activity of PHF2 in vivo, or perhaps PHF2 acts on non-histone substrates. Furthermore, PHF2 shares significant sequence homolog throughout the entire region, including the above-mentioned tyrosine at the corresponding iron-binding position, with that of Schizosaccharomyces pombe Epe1, which plays essential role in heterochromatin function but also has no known enzymatic activity.