Specific labeling of proteins within living cells with single residue precision through genetically encoded click chemistry

The development of catalyst-free click chemistry, i.e. a chemical reaction that is non-toxic and proceeds readily and specifically even within living organisms, has revolutionized chemical biology. By encoding strained alkyne unnatural amino acids into the genetic repertoire of the cell, we exploited this chemistry to label proteins with single residue specificity in bacteria. Meanwhile, we extended the method to more click type reactions, creating a unique opportunity to establish these techniques for installing probes to control and visualize protein function within living species.

In this proposal, we aim to lay the foundation for a smaller and more stable substitute to fluorescent proteins in biology, medicine, and diagnostics. In particular, we aim to explore the potential of click chemistries of the Huisgen-type and inverse electron-demand Diels-Alder reaction and optimize and characterize probes in vitro and in vivo. We will use the techniques to study growth factor receptor signaling and to develop genetically encoded small molecule dye FRET sensors

Dr. Edward Lemke
European Molecular Biology Laboratory (EMBL)

Tel.: +49 6221 387 - 8526

Email Dr. Lemke

Dr. Carsten Schultz
European Molecular Biology Laboratory (EMBL)

Email Dr. Schultz

Publications within the SPP 1623 project

P. Werther+, K. Yserentant+, F. Braun, K. Grussmayer, V. Navikas, M. Yu, Z. Zhang, M.J. Ziegler, C. Mayer, A.J. Gralak, M. Busch, W. Chi, F. Rominger, A. Radenovic, X. Liu, E.A. Lemke, T. Buckup, D.-P. Herten*, R. Wombacher*
ACS Cent. Sci. 2021, 7(9), 1561–157
Bioorthogonal red and far-red fluorogenic probes for wash-free live-cell and super-resolution microscopy
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C.D. Reinkemeier, C. Koehler, P.F. Sauter, N.V. Shymanska, C. Echalier, A. Rutkowska, D.W. Will, C. Schultz, E.A. Lemke
Chemistry2021, 27(19), 6094-6099
Synthesis and Evaluation of Novel Ring-Strained Noncanonical Amino Acids for Residue-Specific Bioorthogonal Reactions in Living Cells
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C. Koehler, G. Estrada Girona, C.D. Reinkemeier, E.A. Lemke
Chembiochem2020, 21(22), 3216-3219
Inducible Genetic Code Expansion in Eukaryotes
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C.D. Reinkemeier, G. Estrada Girona, E.A. Lemke
Science2019, 363, No. 6434
Designer membraneless organelles enable codon reassignment of selected mRNAs in eukaryotes
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A. Kormos, C. Koehler, E.A. Fodor, Z.R. Rutkai, M.E. Martin, G. Mező, E.A. Lemke, P. Kele
Chemistry 2018, 24(35), 8841-8847
Bistetrazine-Cyanines as Double-Clicking Fluorogenic Two-Point Binder or Crosslinker Probes
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E. Kozma, G. Estrada Girona, G. Paci, E.A. Lemke, P. Kele
Chem Commun (Camb) 2017, 53(50), 6696-6699
Bioorthogonal double-fluorogenic siliconrhodamine probes for intracellular super-resolution microscopy
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V. Sakin, J. Hanne, J. Dunder, M. Anders-Össwein, V. Laketa, I. Nikic, H.G. Kräusslich, E.A. Lemke, B. Müller
Cell Chem Biol. 2017, 24(5), 635-645
A Versatile Tool for Live-Cell Imaging and Super-Resolution Nanoscopy Studies of HIV-1 Env Distribution and Mobility
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O.Demeter, A. Kormos, C. Koehler, G. Mezo, K. Németh, E. Kozma, L.B. Takács, E.A. Lemke, P. Kele
Bioconjug Chem. 2017, 28(5), 1552-1559
Bisazide Cyanine Dyes as Fluorogenic Probes for Bis-Cyclooctynylated Peptide Tags and as Fluorogenic Cross-Linkers of Cyclooctynylated Proteins.
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I. Nikic, G.E. Girona, J.H. Kang, G. Paci, S. Mikhaleva, C. Koehler, N. V. Shymanska, C.V. Santos, D. Spitz, E.A. Lemke
Angew. Chem. Int. Ed. 2016, 55(52), 16172-16176
Debugging Eukaryotic Genetic Code Expansion for Site-Specific Click-PAINT Super-Resolution Microscopy
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B. Simon, A.-S. Huart, K. Temmerman, J. Vahokoski, H.D.T. Mertens, D. Komadina, J.-E. Hoffmann, H. Yumerefendi, D. Svergun, P. Kursuli, C. Schultz, D. Hart, A. McCarthy, M. Wilmanns
Structure 2016, 24, 851-861.
Death-associated protein kinase activity is regulated by coupled calcium/calmodulin binding to two distinct sites.
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C. Koehler, P. Sauter, M. Wawryszyn, G. Estrada Girona, K. Gupta, J.J.M. Landry, M.H. Fritz, K. Radic, J.-E. Hoffmann, Z. Angel Chen, J. Zou, P. Siong Tan, B. Galik, S. Junttila, P. Stolt-Bergner, G. Pruneri, A. Gyenesei, C. Schultz, M. Bosse Biskup, H. Besir, V. Benes, J. Rappsilber, M. Jechlinger, J.O. Korbel, I Berger, S. Braese, E.A. Lemke
Nat. Methods 2016, 13, 997-1000
Genetic code expansion for multiprotein complex engineering
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J.A. Wagner, D. Mercadante, I. Nikic, E.A. Lemke, F. Gräter
Chem. Eur. J. 2015 , 21(35), 12431-12435
Orthogonality of Strain-Promoted Click Reactions
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J.E. Hoffmann, T. Plass, I. Nikic, I.V. Aramburu, C. Köhler, H. Gillandt, E.A. Lemke, C. Schultz
Chemistry 2015, 21(35), 12266-12270
Highly stable trans-cyclooctene amino acids for live cell labeling
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S. Tyagi, E.A. Lemke
Curr. Opin. Struct. Biol. 2015, 32, 66-73
Single-molecule FRET and crosslinking studies in structural biology enabled by noncanonical amino acids.
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I. Nikic, T. Plass, O. Schraidt, J. Szymanski, J.A. Briggs, C. Schultz, E.A. Lemke
Angew. Chem. Int. Ed.2014, 53 (8), 2245-2249
Minimal Tags for Rapid Dual-Color Live-Cell Labeling and Super-Resolution Microscopy
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E.A. Lemke
Chembiochem. 2014, 15(12), 1691-1694
The exploding genetic code.
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A. Herner, I. Nikic, M. Kallay, E.A. Lemke, P. Kele
Org Biomol Chem 2013, 11(20), 3297-3306
A new family of bioorthogonally applicable fluorogenic labels.
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S. Milles, E.A. Lemke
Bioessays2013, 35(1), 65-74
What precision-protein-tuning and nano-resolved single molecule sciences can do for each other
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