Abstract |
The histone methyltransferase MLL1 has been linked to translocation-associated gene fusion in childhood leukemias and is an attractive drug target. High-throughput biochemical analysis of MLL1 methyltransferase activity requires the production of at least a trimeric complex of MLL1, RbBP5 and WDR5 to elicit robust activity. Production of trimeric and higher order MLL1 complexes in the quantities and reproducibility required for high-throughput screening presents a significant impediment to MLL1 drug discovery efforts. We present here a small molecule fluorescent ligand (FL-NAH, 6) that is able to bind to the S-adenosylmethionine (SAM) binding site of MLL1 in a manner independent of the associated complex members. We have used FL-NAH to develop a fluorescence polarization-based SAM displacement assay in a 384-well format targeting the MLL1 SET domain in the absence of associated complex members. FL-NAH competes with SAM and is displaced from the MLL1 SET domain by other SAM-binding site ligands with Kdisp values similar to the higher-order complexes, but is unaffected by the H3 peptide substrate. This assay enables screening for SAM-competitive MLL1 inhibitors without requiring the use of trimeric or higher order MLL1 complexes, significantly reducing screening time and cost.
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Authors | Yepeng Luan, Levi L Blazer, Hao Hu, Taraneh Hajian, Jing Zhang, Hong Wu, Scott Houliston, Cheryl H Arrowsmith, Masoud Vedadi, Yujun George Zheng |
Journal | Organic & biomolecular chemistry
(Org Biomol Chem)
Vol. 14
Issue 2
Pg. 631-638
(Jan 14 2016)
ISSN: 1477-0539 [Electronic] England |
PMID | 26541578
(Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, Non-P.H.S.)
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Chemical References |
- Enzyme Inhibitors
- KMT2A protein, human
- Ligands
- Small Molecule Libraries
- Myeloid-Lymphoid Leukemia Protein
- S-Adenosylmethionine
- Histone-Lysine N-Methyltransferase
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Topics |
- Binding, Competitive
(drug effects)
- Dose-Response Relationship, Drug
- Drug Design
- Drug Evaluation, Preclinical
(economics, methods)
- Enzyme Inhibitors
(chemical synthesis, chemistry, pharmacology)
- Fluorescence
- Histone-Lysine N-Methyltransferase
(chemistry, metabolism)
- Humans
- Ligands
- Molecular Structure
- Myeloid-Lymphoid Leukemia Protein
(chemistry, metabolism)
- Protein Structure, Tertiary
- S-Adenosylmethionine
(metabolism)
- Small Molecule Libraries
(chemical synthesis, chemistry, pharmacology)
- Structure-Activity Relationship
- Time Factors
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