Methylation at the 5 position of the cytosine base, when followed by guanine (CpG) in the promoter region of a protein-coding gene, is an epigenetic modification that has been shown to be involved in gene silencing. Methyl binding domain (MBD) family proteins specifically bind double-stranded, methylated DNA which makes them excellent transducers for DNA methylation in biosensing applications. Using yeast surface display, I engineered an MBD variant with improved binding affinity, concatenated it, and expressed it in E. coli as a green fluorescent protein (GFP) fusion. Using these MBD-GFP fusion proteins, I developed a simple method for detecting methylated DNA following direct hybridization on a biochip with either fluorescent or colorimetric (visible hydrogel) readout formed by radical photopolymerization with short (<2 min) reaction times.
Brandon W. Heimer, Brooke Tam, Alissa F. Minkovsky and Hadley D. Sikes. “Using nanobiotechnology to increase the prevalence of epigenotyping assays in precision medicine,” WIREs Nanomedicine & Nanobiotechnology, 2017, 9 (1). DOI: 10.1002/wnan.1407.
Brandon W. Heimer, Brooke E. Tam and Hadley D. Sikes. “Characterization and Directed Evolution of a Methyl Binding Domain Protein for High-Sensitivity DNA Methylation Analysis,” Protein Engineering, Design and Selection, 2015. DOI:10.1093/protein/gzv046.
B.W. Heimer, T.A. Shatova, J.K. Lee, K. Kaastrup and H.D. Sikes. “Evaluating the Sensitivity of Hybridization-Based Epigenotyping using a Methyl Binding Domain Protein,” Analyst, 2014, 139 (15): 3695-3701. DOI:10.1039/C4AN00667D. (Cover Story)
J.K. Lee, B.W. Heimer, H.D. Sikes. “Systematic study of fluorescein-functionalized macrophotoinitiators for colorimetric bioassays,” Biomacromolecules, 2012, 13 (4): 1136–1143. DOI:10.1021/bm300037t.
M.E. Boyd, B.W. Heimer, H.D. Sikes. “Functional heterologous expression and purification of a mammalian methyl-CpG binding domain in suitable yield for DNA methylation profiling assays,” Protein Expression & Purification, 2012, 82 (2): 332-338. DOI:10.1016/j.pep.2012.01.016.
Staff scientist/engineer, Sandia National Laboratories, Livermore