title: Disruption of Bcr-Abl coiled coil oligomerization by design.
abstract: Oligomerization is an important regulatory mechanism for many proteins, including oncoproteins and other pathogenic proteins. The oncoprotein Bcr-Abl relies on oligomerization via its coiled coil domain for its kinase activity, suggesting that a designed coiled coil domain with enhanced binding to Bcr-Abl and reduced self-oligomerization would be therapeutically useful. Key mutations in the coiled coil domain of Bcr-Abl were identified that reduce homo-oligomerization through intermolecular charge-charge repulsion yet increase interaction with the Bcr-Abl coiled coil through additional salt bridges, resulting in an enhanced ability to disrupt the oligomeric state of Bcr-Abl. The mutations were modeled computationally to optimize the design. Assays performed in vitro confirmed the validity and functionality of the optimal mutations, which were found to exhibit reduced homo-oligomerization and increased binding to the Bcr-Abl coiled coil domain. Introduction of the mutant coiled coil into K562 cells resulted in decreased phosphorylation of Bcr-Abl, reduced cell proliferation, and increased caspase-3/7 activity and DNA segmentation. Importantly, the mutant coiled coil domain was more efficacious than the wild type in all experiments performed. The improved inhibition of Bcr-Abl through oligomeric disruption resulting from this modified coiled coil domain represents a viable alternative to small molecule inhibitors for therapeutic intervention.
authors.affiliation: Department of Pharmaceutics and Pharmaceutical Chemistry, College of Pharmacy, University of Utah, Salt Lake City, Utah 84108, USA.
authors.forename: Andrew S
authors.initials: AS
authors.lastname: Dixon
authors.forename: Scott S
authors.initials: SS
authors.lastname: Pendley
authors.forename: Benjamin J
authors.initials: BJ
authors.lastname: Bruno
authors.forename: David W
authors.initials: DW
authors.lastname: Woessner
authors.forename: Adrian A
authors.initials: AA
authors.lastname: Shimpi
authors.forename: Thomas E
authors.initials: TE
authors.lastname: Cheatham
authors.forename: Carol S
authors.initials: CS
authors.lastname: Lim
mesh_terms: D000818:Animals; D001704:Biopolymers / Q000737:chemistry*; D015153:Blotting, Western; D019556:COS Cells; D049109:Cell Proliferation; D002522:Chlorocebus aethiops; D003198:Computer Simulation; D016044:Fusion Proteins, bcr-abl / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D006860:Hydrogen Bonding; D020014:K562 Cells; D008856:Microscopy, Fluorescence; D008958:Models, Molecular; D016296:Mutagenesis; D010957:Plasmids; D011485:Protein Binding; D013816:Thermodynamics; D020798:Two-Hybrid System Techniques
chemical_list: D001704:Biopolymers; D016044:Fusion Proteins, bcr-abl