Optimizing methodology for the detection of H3K27me3 levels using flow cytometry

<p>Optimizing methods to detect H3K27m3 levels using flow cytometry in patient AML cells.</p

Determining the plasticity of CD34 expression

<p>Determining if OCI-AML-20 cells can regain CD34 expression.</p

Mass Spectrometry of Huntingtin Fragments (2016/02/06)

<p>Open lab notebook write up for project: huntingtin structural studies.</p

Mass Spectrometry .raw Files and Methods for Huntingtin Limited Proteolysis (2016/02/11)

<p>Open lab notebook for project: huntingtin structural studies</p

Structural and Functional Survey of Environmental Aminoglycoside Acetyltransferases Reveals Functionality of Resistance Enzymes

Aminoglycoside <i>N</i>-acetyltransferases (AACs) confer resistance against the clinical use of aminoglycoside antibiotics. The origin of AACs can be traced to environmental microbial species representing a vast reservoir for new and emerging resistance enzymes, which are currently undercharacterized. Here, we performed detailed structural characterization and functional analyses of four metagenomic AAC (meta-AACs) enzymes recently identified in a survey of agricultural and grassland soil microbiomes (Forsberg et al. Nature 2014, 509, 612). These enzymes are new members of the Gcn5-Related-<i>N</i>-Acetyltransferase superfamily and confer resistance to the aminoglycosides gentamicin C, sisomicin, and tobramycin. Moreover, the meta-AAC0020 enzyme demonstrated activity comparable with an AAC(3)-I enzyme that serves as a model AAC enzyme identified in a clinical bacterial isolate. The crystal structure of meta-AAC0020 in complex with sisomicin confirmed an unexpected AAC(6′) regiospecificity of this enzyme and revealed a drug binding mechanism distinct from previously characterized AAC(6′) enzymes. Together, our data highlights the presence of highly active antibiotic-modifying enzymes in the environmental microbiome and reveals unexpected diversity in substrate specificity. These observations of additional AAC enzymes must be considered in the search for novel aminoglycosides less prone to resistance

Discovery of a Potent Class I Protein Arginine Methyltransferase Fragment Inhibitor

Protein methyltransferases (PMTs) are a promising target class in oncology and other disease areas. They are composed of SET domain methyltransferases and structurally unrelated Rossman-fold enzymes that include protein arginine methyltransferases (PRMTs). In the absence of a well-defined medicinal chemistry tool-kit focused on PMTs, most current inhibitors were identified by screening large and diverse libraries of leadlike molecules. So far, no successful fragment-based approach was reported against this target class. Here, by deconstructing potent PRMT inhibitors, we find that chemical moieties occupying the substrate arginine-binding site can act as efficient fragment inhibitors. Screening a fragment library against PRMT6 produced numerous hits, including a 300 nM inhibitor (ligand efficiency of 0.56) that decreased global histone 3 arginine 2 methylation in cells, and can serve as a warhead for the development of PRMT chemical probes