Protocol for Biochemical Analysis and Structure Determination of the ZZ Domain of the E3 Ubiquitin Ligase HERC2

STAR Protoc. 2020 Oct 28;1(3):100155. doi: 10.1016/j.xpro.2020.100155. eCollection 2020 Dec 18.

Abstract

Since its discovery, several ligands of the ZZ domain have been identified; however, molecular and structural information underlying binding of these ligands remains limited. Here, we describe a protocol for biochemical and structural analysis of the ZZ domain of human E3 ubiquitin ligase HERC2 (HERC2ZZ) and its interaction with its ligands: the N-terminal tails of histone H3 and SUMO1. This methodology could be applied for characterization of binding activities of other histone readers. For complete details on the use and execution of this protocol, please refer to Liu et al. (2020).

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biochemistry / methods*
  • Buffers
  • Crystallization
  • Fluorescence
  • HEK293 Cells
  • Histones / metabolism
  • Humans
  • Magnetic Resonance Spectroscopy
  • Peptides / metabolism
  • Protein Domains
  • Recombinant Fusion Proteins / isolation & purification
  • SUMO-1 Protein / isolation & purification
  • SUMO-1 Protein / metabolism
  • Tryptophan / metabolism
  • Ubiquitin-Protein Ligases / chemistry*
  • Ubiquitin-Protein Ligases / metabolism

Substances

  • Buffers
  • Histones
  • Peptides
  • Recombinant Fusion Proteins
  • SUMO-1 Protein
  • SUMO1 protein, human
  • Tryptophan
  • HERC2 protein, human
  • Ubiquitin-Protein Ligases