Structural and Functional Role of Acetyltransferase hMOF K274 Autoacetylation

Cheryl E. McCullough; Shufei Song; Michael H. Shin; F. Brad Johnson; Ronen Marmorstein

doi:10.1074/jbc.M116.736264

Structural and Functional Role of Acetyltransferase hMOF K274 Autoacetylation*

From the ^‡Department of Biochemistry and Biophysics, Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104,
the ^§Department of Chemistry, University of Pennsylvania, Philadelphia, Pennsylvania 19104, and
the ^¶Department of Pathology and Laboratory Medicine and
^‖Graduate Group in Biochemistry and Molecular Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104

↵1 To whom correspondence should be addressed: Dept. of Biochemistry and Biophysics, Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, 421 Curie Blvd., Philadelphia, PA 19104. Tel.: 215-898-7740; Fax: 215-746-5511; E-mail: marmor{at}mail.med.upenn.edu.

Abstract

Many histone acetyltransferases undergo autoacetylation, either through chemical or enzymatic means, to potentiate enzymatic cognate substrate lysine acetylation, although the mode and molecular role of such autoacetylation is poorly understood. The MYST family of histone acetyltransferases is autoacetylated at an active site lysine residue to facilitate cognate substrate lysine binding and acetylation. Here, we report on a detailed molecular investigation of Lys-274 autoacetylation of the human MYST protein Males Absent on the First (hMOF). A mutational scan of hMOF Lys-274 reveals that all amino acid substitutions of this residue are able to bind cofactor but are significantly destabilized, both in vitro and in cells, and are catalytically inactive for cognate histone H4 peptide lysine acetylation. The x-ray crystal structure of a hMOF K274P mutant suggests that the reduced stability and catalytic activity stems from a disordering of the residue 274-harboring a α2-β7 loop. We also provide structural evidence that a C316S/E350Q mutant, which is defective for cognate substrate lysine acetylation; and biochemical evidence that a K268M mutant, which is defective for Lys-274 chemical acetylation in the context of a K274-peptide, can still undergo quantitative K274 autoacetylation. Together, these studies point to the critical and specific role of hMOF Lys-274 autoacetylation in hMOF stability and cognate substrate acetylation and argues that binding of Ac-CoA to hMOF likely drives Lys-274 autoacetylation for subsequent cognate substrate acetylation.

Footnotes

↵* This work was supported, in whole or in part, by National Institutes of Health Grants R01 GM060293 and P01 AG031862 (to R. M. and B. J.) and T32 GM008275 (to C. E. M.), and a predoctoral fellowship grant from the American Heart Association (to S. S.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
The atomic coordinates and structure factors (codes 5J8F and 5J8C) have been deposited in the Protein Data Bank (http://wwpdb.org/).