Journal
ACS CHEMICAL BIOLOGY
Volume 10, Issue 8, Pages 1880-1886Publisher
AMER CHEMICAL SOC
DOI: 10.1021/acschembio.5b00201
Keywords
-
Categories
Funding
- National Institutes of Health [R01GM107520]
- Lou Hegedus Fellowship
Ask authors/readers for more resources
Overexpression of the ankyrin repeat oncoprotein gankyrin is directly linked to the onset, proliferation, and/or metastasis of many cancers. The role of gankyrin in multiple disease-relevant biochemical processes is profound. In addition to other cellular processes, gankyrin overexpression leads to decreased cellular levels of p53, through a complex that involves MDM2. Thus, inhibition of this interaction is an attractive strategy for modulating oncogenic phenotypes in gankyrin-overexpressing cells. However, the lack of well-defined, hydrophobic, small-molecule binding pockets on the putative ankyrin repeat binding face presents a challenge to traditional small-molecule drug discovery. In contrast, by virtue of their size and relatively high folding energies, synthetic gankyrin-binding proteins could, in principle, compete with physiologically relevant PPIs involving gankyrin. Previously, we showed that a shape-complementary protein scaffold can be resurfaced to bind gankyrin with moderate affinity (K-D similar to 6 mu M). Here, we used yeast display high-throughput screening, error-prone PCR, DNA shuffling, and protein engineering to optimize proteins bind gankyrin with excellent affinity (K-D similar to 21 nM), selectively co-purifies with gankyrin from a complex cellular milieu, modulates an interaction between gankyrin and a physiological binding partner (S6 ATPase), and suppresses gankyrin/MDM2-dependent ubiquitination of p53.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available