Genetically Encoded Chemical Decaging in Living Bacteria

Lu, Lingbin; Liu, Yanjun; Zhang, Gong; Ge, Yun; Fan, Xinyuan; Feng, Lin; Wang, Jie; Zheng, Huangtao; Xie, Xiao; Zeng, Xiangmei; Chen, Peng R.

doi:10.1021/acs.biochem.7b01017

Cited by 33 publications

(32 citation statements)

References 34 publications

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“…9,[13][14][15][16][17] These reactions are particularly useful for precision protein labeling, decaging, and cellular protein imaging. 13,14,[17][18][19][20][21][22][23][24] For example, tetrazine-containing amino acids have been genetically incorporated into proteins, allowing for a rapid bioorthogonal conjugation with trans-cyclooctene (TCO)labeled biomolecules. 13,17 Taking advantage of the high reaction rate and good biocompatibility of IEDDA reactions, Chen and others have redirected this reaction for sitespecific protein decaging.…”

Section: Graphical Abstractmentioning

confidence: 99%

Addition of Isocyanide-Containing Amino Acids to the Genetic Code for Protein Labeling and Activation

Chen

Tang

et al. 2019

ACS Chem. Biol.

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Site-specific introduction of bioorthogonal handles into biomolecules provides powerful tools for studying and manipulating the structures and functions of proteins. Recent advances in bioorthogonal chemistry demonstrate that tetrazine-based bioorthogonal cycloaddition is a particularly useful methodology due to its high reactivity, biological selectivity, and turn-on property for fluorescence imaging. Despite its broad applications in protein labeling and imaging, utilization of tetrazine-based bioorthogonal cycloaddition has been limited to date by the requirement of a hydrophobic strained alkene reactive moiety. Circumventing this structural requirement, we report the site-specific incorporation of noncanonical amino acids (ncAAs) with a small isocyanide (or isonitrile) group into proteins in both bacterial and mammalian cells. We showed that under physiological conditions and in the absence of a catalyst these isocyanidecontaining ncAAs could react selectively with tetrazine molecules via [4 + 1]-cycloaddition, thus providing a versatile bioorthogonal handle for site-specific protein labeling and protein decaging. Significantly, these bioorthogonal reactions between isocyanides and tetrazines also provide a unique mechanism for the activation of tetrazine-quenched fluorophores. The addition of these isocyanide-containing ncAAs to the list of 20 commonly used, naturally occurring amino acids expands our repertoire of reagents for bioorthogonal chemistry, therefore enabling new biological applications ranging from protein labeling and imaging studies to the chemical activation of proteins.

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Section: Graphical Abstractmentioning

confidence: 99%

Addition of Isocyanide-Containing Amino Acids to the Genetic Code for Protein Labeling and Activation

Chen

Tang

et al. 2019

ACS Chem. Biol.

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“…Subsequently, we analyzed whether deprotection with the prokaryote-compatible 3,6-dimethyl-tetrazine (26) could be used to switch on recombinant protein expression. 12,42 We found that 26 caused a minor delay on IPG (15) induced OVA expression without reducing overall expression levels ( Fig. S5 and Table S10 †).…”

Section: Resultsmentioning

confidence: 94%

Synthetic methodology towards allylictrans-cyclooctene-ethers enables modification of carbohydrates: bioorthogonal manipulation of thelacrepressor

et al. 2020

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“…Similarly, “Syn‐notch” receptors bear an antigen recognition domain and signal through the Notch pathway . Finally, a plethora of chimeric proteins have been engineered that combine fluorescent proteins with sensory domains and have been used to quantify endogenous small molecules . However, to the best of our knowledge, synthetic receptors have never been created by combining protein modules from GPCRs and RTKs.…”

Section: Figurementioning

confidence: 99%

Transformation of Receptor Tyrosine Kinases into Glutamate Receptors and Photoreceptors

et al. 2020

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Receptor tyrosine kinases (RTKs) are key regulators of cellular functions in metazoans. In vertebrates, RTKs are mostly activated by polypeptides but are not naturally sensitive to amino acids or light. Taking inspiration from Venus kinase receptors (VKRs), an atypical family of RTKs found in nature, we have transformed the human insulin (hIR) and hepatocyte growth factor receptor (hMET) into glutamate receptors by replacing their extracellular binding domains with the ligand‐binding domain of metabotropic glutamate receptor type 2 (mGluR2). We then imparted light sensitivity through covalent attachment of a synthetic glutamate‐based photoswitch via a self‐labelling SNAP tag. By employing a Xenopus laevis oocyte kinase activity assay, we demonstrate how these chimeric RTKs, termed light‐controlled human insulin receptor (LihIR) and light‐controlled human MET receptor (LihMET), can be used to exert optical control over the insulin or MET signaling pathways. Our results outline a potentially general strategy to convert RTKs into photoreceptors.

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Genetically Encoded Chemical Decaging in Living Bacteria

Cited by 33 publications

References 34 publications

Addition of Isocyanide-Containing Amino Acids to the Genetic Code for Protein Labeling and Activation

Addition of Isocyanide-Containing Amino Acids to the Genetic Code for Protein Labeling and Activation

Synthetic methodology towards allylictrans-cyclooctene-ethers enables modification of carbohydrates: bioorthogonal manipulation of thelacrepressor

Transformation of Receptor Tyrosine Kinases into Glutamate Receptors and Photoreceptors

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