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Methods Enzymol. 2016;573:321-43. doi: 10.1016/bs.mie.2016.01.018. Epub 2016 Mar 24.

Preparation, Biochemical Analysis, and Structure Determination of the Bromodomain, an Acetyl-Lysine Binding Domain.

Author information

1
Icahn School of Medicine at Mount Sinai, New York, NY, United States.
2
Icahn School of Medicine at Mount Sinai, New York, NY, United States. Electronic address: ming-ming.zhou@mssm.edu.

Abstract

The bromodomain (BrD) represents an evolutionarily conserved protein domain whose function mostly is to recognize acetylated lysine residues in histones and nuclear proteins in regulation of gene transcription in chromatin. The highly conserved BrD structure features an unusual left-handed, antiparallel four-helix bundle and a hydrophobic pocket between the interhelical ZA and BC loops important for acetyl-lysine binding. Many proteins, particularly transcriptional activators, contain BrDs, and mutation or deletion of the BrDs impairs the protein function, implying their critical role in human biology and disease. In this chapter, we provide general protocols of the preparation, biochemical analysis, and structure determination of BrDs, aiming to offer a general guideline for structural and biochemical functional characterization of BrD-containing proteins.

KEYWORDS:

Acetyl-lysine binding; Bromodomain; Chromatin biology; Gene transcription; Histone lysine acetylation; NMR spectroscopy; Protein purification and characterization; Structural biology

PMID:
27372760
DOI:
10.1016/bs.mie.2016.01.018
[Indexed for MEDLINE]

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