posted on 2022-12-29, 18:34authored byWeijun Su, Yi Wang, Siqi Zou, Yanjie Zhao, Yifan Li, Chunze Zhang, Xiaojing Guo, Shuai Li
While different display
technologies, represented by
phage display,
have been widely used in drug discovery, they still can hardly achieve
function-based peptide screening, which in most cases is performed
in mammalian cells. And most attempts to screen functional peptides
with mammalian platforms utilized plasmids to store coding information.
Our previous work established double-stranded DNAs (dsDNAs) as innovative
biological parts to implement AND-gate genetic circuits in mammalian
cells. In the current study, we employ dsDNAs with terminal NNK degenerate
codons to implement AND-gate genetic circuits and generate peptide
libraries in mammalian cells. This dsDNA-based AND-gate (DBAG) peptide
library construction strategy is easy to perform, requiring only PCR
reaction and cell transfection. High-throughput sequencing (HTS) and
single-cell sequencing results revealed both peptide length and amino
acid sequence diversity of DBAG peptide libraries. Moreover, as a
feasibility test of this strategy, we identified an MDM2-interacting
peptide by applying the DBAG peptide library to a mammalian cell-based
two-hybrid system. Our work establishes dsDNAs with terminal degenerate
codons as biological parts to build peptide libraries in mammalian
cells, which may have great application potential in the future.