Abstract
We describe the use of iterative in situ click chemistry to design an Akt-specific branched peptide triligand that is a drop-in replacement for monoclonal antibodies in multiple biochemical assays. Each peptide module in the branched structure makes unique contributions to affinity and/or specificity resulting in a 200 nM affinity ligand that efficiently immunoprecipitates Akt from cancer cell lysates and labels Akt in fixed cells. Our use of a small molecule to preinhibit Akt prior to screening resulted in low micromolar inhibitory potency and an allosteric mode of inhibition, which is evidenced through a series of competitive enzyme kinetic assays. To demonstrate the efficiency and selectivity of the protein-templated in situ click reaction, we developed a novel QPCR-based methodology that enabled a quantitative assessment of its yield. These results point to the potential for iterative in situ click chemistry to generate potent, synthetically accessible antibody replacements with novel inhibitory properties.
Original language | English (US) |
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Pages (from-to) | 18280-18288 |
Number of pages | 9 |
Journal | Journal of the American Chemical Society |
Volume | 133 |
Issue number | 45 |
DOIs | |
State | Published - Nov 16 2011 |
Externally published | Yes |
ASJC Scopus subject areas
- Catalysis
- General Chemistry
- Biochemistry
- Colloid and Surface Chemistry