Engineered Reader Proteins for Enhanced Detection of Methylated Lysine on Histones.

Histone post-translational modifications (PTMs) are crucial for many cellular processes including mitosis, transcription, and DNA repair. Accordingly, methods that report on the presence of PTMs are essential tools for resolving open questions about epigenetic processes and for developing therapeutic diagnostics. Reader domains that recognize histone PTMs have shown potential as advantageous substitutes for anti-PTM antibodies and engineering efforts aimed at enhancing reader domain affinities would advance their efficacy as antibody alternatives. Here, we describe engineered chromodomains from D. melanogaster and humans that bind more tightly to H3K9 methylation (H3K9me) marks and result in the tightest reported reader:H3K9me interaction to date. Furthermore, we show that these engineered readers (eReaders) increase detection of H3K9me marks in several biochemical assays and outperform a commercial anti-H3K9me antibody in detecting H3K9me-containing nucleosomes in vitro, demonstrating the utility of eReaders to complement antibodies in epigenetics research.