title: Optimized Tetrazine Derivatives for Rapid Bioorthogonal Decaging in Living Cells.
abstract: The inverse-electron-demand Diels-Alder (iDA) reaction has recently been repurposed as a bioorthogonal decaging reaction by accelerating the elimination process after an initial cycloaddition between trans-cyclooctene (TCO) and tetrazine (TZ). Herein, we systematically surveyed 3,6-substituted TZ derivatives by using a fluorogenic TCO-coumarin reporter followed by LC-MS analysis, which revealed that the initial iDA cycloaddition step was greatly accelerated by electron-withdrawing groups (EWGs) while the subsequent elimination step was strongly suppressed by EWGs. In addition, smaller substituents facilitated the decaging process. These findings promoted us to design and test unsymmetric TZs bearing an EWG group and a small non-EWG group at the 3- and 6-position, respectively. These TZs showed remarkably enhanced decaging rates, enabling rapid iDA-mediated protein activation in living cells.
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871, China.|Peking-Tsinghua Center for Life Sciences, Beijing, 100871, China.
authors.forename: Xinyuan
authors.initials: X
authors.lastname: Fan
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871, China.
authors.forename: Yun
authors.initials: Y
authors.lastname: Ge
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871, China.
authors.forename: Feng
authors.initials: F
authors.lastname: Lin
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871, China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871, China.|Peking-Tsinghua Center for Life Sciences, Beijing, 100871, China.
authors.forename: Gong
authors.initials: G
authors.lastname: Zhang
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871, China.
authors.forename: William Shu Ching
authors.initials: WS
authors.lastname: Ngai
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871, China.
authors.forename: Zhi
authors.initials: Z
authors.lastname: Lin
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871, China.
authors.forename: Siqi
authors.initials: S
authors.lastname: Zheng
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871, China.|Peking-Tsinghua Center for Life Sciences, Beijing, 100871, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Wang
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871, China.|Peking-Tsinghua Center for Life Sciences, Beijing, 100871, China.
authors.forename: Jingyi
authors.initials: J
authors.lastname: Zhao
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Li
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871, China. pengchen@pku.edu.cn.|Peking-Tsinghua Center for Life Sciences, Beijing, 100871, China. pengchen@pku.edu.cn.
authors.forename: Peng R
authors.initials: PR
authors.lastname: Chen
mesh_terms: D003374:Coumarins / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000737:chemistry*; D049452:Green Fluorescent Proteins / Q000737:chemistry; D057809:HEK293 Cells; D006801:Humans; D013777:Tetrazoles / Q000032:analysis* / Q000378:metabolism
chemical_list: D003374:Coumarins; D005456:Fluorescent Dyes; D013777:Tetrazoles; D049452:Green Fluorescent Proteins; C030123:coumarin
keywords: Diels-Alder reactions; amino acids; decaging; protein activation; protein engineering