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Red and far-red cleavable fluorescent dyes for self-labelling enzyme protein tagging

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Winners: 2024 Outstanding Early Career Research Award of 蜜桃社区 Chemical Biology

Johannes Broichhagen, Kilian Roßmann, Ramona Birke, Joshua Levitz and Ben Jones

Paper: Red and far-red cleavable fluorescent dyes for self-labelling enzyme protein tagging and interrogation of GPCR co-internalization

G protein-coupled receptors (GPCRs) are transmembrane receptors which serve as major drug targets for a plethora of diseases. Understanding their intricate signaling and trafficking properties, including signal termination by endocytosis, is at the forefront of modern research in this field.

In our study, we developed post-labelling cleavable substrates for self-labelling protein tags to study cell surface receptor trafficking events by stripping dyes from non-internalized protein pools. We report the development of four membrane impermeable probes that covalently bind to either the SNAP- or the Halo-tag in the red to far-red color spectrum, and can be removed exclusively from the surface using the reducing agent sodium 2-mercaptoethane sulfonate (MESNA). We use this approach to detect internalized homo- and heterodimers of metabotropic glutamate receptors (mGluRs) and reveal unidirectional crosstalk between co-expressed glucagon-like peptide 1 (GLP1R) and glucose-dependent insulinotropic polypeptide receptors (GIPR).

In these applications, we translate our method to both high resolution imaging and quantitative, high throughput assays, demonstrating the value of our approach for a wide range of applications, for instance for ligand screening.

About the winners

Johannes_Broichhagen

Johannes studied chemistry at鈥痶he University of Erlangen-Nuremberg including a one-year stint with Marcus Weck at NYU and鈥痝raduated in 2010 with鈥痟onors. He then joined the group of Dirk Trauner at LMU Munich鈥痶o dive into Chemical鈥疊iology and obtained his鈥疨hD鈥痠n 2014. Next, he was working with Kai Johnsson at EPF Lausanne and at the MPI for Medical Research in鈥疕eidelberg. Since March 2020,鈥痟e joined the FMP in Berlin as an independent Junior Group Leader in Chemical Biology. His research interests lie at the interface of boosting photophysics of chromophores chemically for imaging and optical control, as well as studying cell surface proteins in a native context. He has received numerous prestigious grants (e.g. ERC Starting Grant) and has been recognized with multiple awards for his creative and innovative research approach (ORCHEM 2022, GBM 2022).

Kilian_Rossmann

Kilian studied chemistry at the Julius-Maximilian-University W眉rzburg (JMU) with focus鈥痮n Organic, Physical and Medicinal Chemistry. For his Master Thesis he joined the group of Dirk Trauner at New York University (NYU), where he worked on photoswitchable PROTACs. After graduating in 2019, Kilian decided to join JB鈥檚 lab in 2020 to work on novel imaging techniques, graduating with a PhD in 2025.

Ramona_Birke

Ramona studied Biology with a focus on genetics and molecular biology at the Humboldt University Berlin, graduating 2010 in Molecular Ecology. After a stint at WITA GmbH, she joined the FMP as technical assistant in 2011, working in the field Molecular Cellular Physiology and Mass Spectrometry. She teamed up with JB in 2020 to support and pursue quests in Chemical Biology. 

Joshua_Levitz

Joshua received a B.S. in Biology and a B.S. in Physics from American University in 2009. While an undergraduate he worked in the lab of Miguel Holmgren at the National Institutes of Health. As a PhD student and postdoctoral fellow in the lab of Ehud Isacoff at UC Berkeley, Josh developed and applied new optical approaches to study the activation mechanisms and neurophysiology of glutamate receptors and ion channels. He joined the department of biochemistry at Weill Cornell Medicine in 2016 and is currently Associate Professor in the Department of Biochemistry.

Ben_Jones

Ben Jones is a Clinical Associate Professor within the Section of Endocrinology and Investigative Medicine, and a Metabolic Medicine physician based in Imperial College Healthcare NHS Trust and North West London Pathology. His research is focussed on the translational pharmacology of G protein coupled receptors in metabolic disease. He trained in medicine at the University of Cambridge and completed his PhD (supervisor: Prof Steve Bloom) in 2017, during which he applied the emerging concept of "biased agonism" to the GLP-1 receptor. After completing clinical training in 2020 he has was awarded the Imperial Post-Doctoral, Post-CCT Fellowship (IPPRF), and subsequently an MRC Clinician Scientist Fellowship in 2023.