Journal
CELL REPORTS
Volume 25, Issue 4, Pages 884-+Publisher
CELL PRESS
DOI: 10.1016/j.celrep.2018.09.090
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Funding
- National Key R&D Program of China [2017YFA0506700]
- Strategic Priority Research Program of Chinese Academy of Sciences [XDB19030000]
- Shanghai Science and Technology Committee [16ZR1441600, 18ZR1446900]
- Excellent Young Scientists Fund from National Natural Science Foundation of China [81622022]
- Recruitment Program for Young Professionals
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Base editors (BEs) are emerging tools used for precision correction or diversifying mutation. It provides a potential way to recreate somatic hypermutations (SHM) for generating high-affinity antibody, which is usually screened from antigen-challenged animal models or synthetic combinatorial libraries. By comparing somatic mutations in the same genomic context, we screened engineered deaminases and CRISPR-deaminase coupling approaches and updated diversifying base editors (DBEs) to generate SHM. The deaminase used in DBEs retains its intrinsic nucleotide preference and mutates cytidines at its preferred motifs. DBE with AID targets the same hotspots as physiological AID does in vivo, while DBE with other deaminases generates distinct mutation profiles from the same DNA substrate. Down-stream DNA repair pathways further diversified the sequence, while Cas9-nickase restricted mutation spreading. Finally, application of DBE in an antibody display system achieved antibody affinity maturation ex vivo. Our findings provide insight of DBE working mechanism and an alternative antibody engineering approach.
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