RRC ID 65479
Author Kashima D, Kawahara M.
Title Evolution of KIPPIS as a versatile platform for evaluating intracellularly functional peptide aptamers.
Journal Sci Rep
Abstract Chimeric proteins have been widely used to evaluate intracellular protein-protein interactions (PPIs) in living cells with various readouts. By combining an interleukin-3-dependent murine cells and chimeric proteins containing a receptor tyrosine kinase c-kit, we previously established a c-kit-based PPI screening (KIPPIS) system to evaluate and select protein binders. In the KIPPIS components, proteins of interest are connected with a chemically inducible helper module and the intracellular domain of the growth-signaling receptor c-kit, which detects PPIs based on cell proliferation as a readout. In this system, proteins of interest can be incorporated into chimeric proteins without any scaffold proteins, which would be advantageous for evaluating interaction between small peptides/domains. To prove this superiority, we apply KIPPIS to 6 peptide aptamer-polypeptide pairs, which are derived from endogenous, synthetic, and viral proteins. Consequently, all of the 6 peptide aptamer-polypeptide interactions are successfully detected by cell proliferation. The detection sensitivity can be modulated in a helper ligand-dependent manner. The assay results of KIPPIS correlate with the activation levels of Src, which is located downstream of c-kit-mediated signal transduction. Control experiments reveal that KIPPIS clearly discriminates interacting aptamers from non-interacting ones. Thus, KIPPIS proves to be a versatile platform for evaluating the binding properties of peptide aptamers.
Volume 11(1)
Pages 11758
Published 2021-6-3
DOI 10.1038/s41598-021-91287-z
PII 10.1038/s41598-021-91287-z
PMID 34083659
Human and Animal Cells Ba/F3(RCB0805)