[{"department":[{"_id":"HaJa"}],"isi":1,"day":"30","oa":1,"issue":"9","quality_controlled":"1","publisher":"Nature Publishing Group","page":"861 - 869","status":"public","date_created":"2018-12-11T11:44:49Z","_id":"137","abstract":[{"text":"Fluorescent sensors are an essential part of the experimental toolbox of the life sciences, where they are used ubiquitously to visualize intra- and extracellular signaling. In the brain, optical neurotransmitter sensors can shed light on temporal and spatial aspects of signal transmission by directly observing, for instance, neurotransmitter release and spread. Here we report the development and application of the first optical sensor for the amino acid glycine, which is both an inhibitory neurotransmitter and a co-agonist of the N-methyl-d-aspartate receptors (NMDARs) involved in synaptic plasticity. Computational design of a glycine-specific binding protein allowed us to produce the optical glycine FRET sensor (GlyFS), which can be used with single and two-photon excitation fluorescence microscopy. We took advantage of this newly developed sensor to test predictions about the uneven spatial distribution of glycine in extracellular space and to demonstrate that extracellular glycine levels are controlled by plasticity-inducing stimuli.","lang":"eng"}],"user_id":"c635000d-4b10-11ee-a964-aac5a93f6ac1","publication_status":"published","author":[{"first_name":"William","last_name":"Zhang","full_name":"Zhang, William"},{"full_name":"Herde, Michel","first_name":"Michel","last_name":"Herde"},{"first_name":"Joshua","last_name":"Mitchell","full_name":"Mitchell, Joshua"},{"last_name":"Whitfield","first_name":"Jason","full_name":"Whitfield, Jason"},{"last_name":"Wulff","first_name":"Andreas","full_name":"Wulff, Andreas"},{"full_name":"Vongsouthi, Vanessa","last_name":"Vongsouthi","first_name":"Vanessa"},{"last_name":"Sanchez Romero","first_name":"Inmaculada","id":"3D9C5D30-F248-11E8-B48F-1D18A9856A87","full_name":"Sanchez Romero, Inmaculada"},{"first_name":"Polina","last_name":"Gulakova","full_name":"Gulakova, Polina"},{"full_name":"Minge, Daniel","first_name":"Daniel","last_name":"Minge"},{"last_name":"Breithausen","first_name":"Björn","full_name":"Breithausen, Björn"},{"first_name":"Susanne","last_name":"Schoch","full_name":"Schoch, Susanne"},{"full_name":"Janovjak, Harald L","id":"33BA6C30-F248-11E8-B48F-1D18A9856A87","first_name":"Harald L","orcid":"0000-0002-8023-9315","last_name":"Janovjak"},{"full_name":"Jackson, Colin","first_name":"Colin","last_name":"Jackson"},{"last_name":"Henneberger","first_name":"Christian","full_name":"Henneberger, Christian"}],"external_id":{"pmid":["30061718 "],"isi":["000442174500013"]},"volume":14,"project":[{"grant_number":"RGY0084/2012","_id":"255BFFFA-B435-11E9-9278-68D0E5697425","name":"In situ real-time imaging of neurotransmitter signaling using designer optical sensors (HFSP Young Investigator)"}],"intvolume":"        14","main_file_link":[{"open_access":"1","url":"https://www.ncbi.nlm.nih.gov/pubmed/30061718"}],"title":"Monitoring hippocampal glycine with the computationally designed optical sensor GlyFS","article_processing_charge":"No","publication":"Nature Chemical Biology","language":[{"iso":"eng"}],"year":"2018","date_updated":"2023-09-13T08:58:05Z","citation":{"mla":"Zhang, William, et al. “Monitoring Hippocampal Glycine with the Computationally Designed Optical Sensor GlyFS.” <i>Nature Chemical Biology</i>, vol. 14, no. 9, Nature Publishing Group, 2018, pp. 861–69, doi:<a href=\"https://doi.org/10.1038/s41589-018-0108-2\">10.1038/s41589-018-0108-2</a>.","ieee":"W. Zhang <i>et al.</i>, “Monitoring hippocampal glycine with the computationally designed optical sensor GlyFS,” <i>Nature Chemical Biology</i>, vol. 14, no. 9. Nature Publishing Group, pp. 861–869, 2018.","ama":"Zhang W, Herde M, Mitchell J, et al. Monitoring hippocampal glycine with the computationally designed optical sensor GlyFS. <i>Nature Chemical Biology</i>. 2018;14(9):861-869. doi:<a href=\"https://doi.org/10.1038/s41589-018-0108-2\">10.1038/s41589-018-0108-2</a>","chicago":"Zhang, William, Michel Herde, Joshua Mitchell, Jason Whitfield, Andreas Wulff, Vanessa Vongsouthi, Inmaculada Sanchez-Romero, et al. “Monitoring Hippocampal Glycine with the Computationally Designed Optical Sensor GlyFS.” <i>Nature Chemical Biology</i>. Nature Publishing Group, 2018. <a href=\"https://doi.org/10.1038/s41589-018-0108-2\">https://doi.org/10.1038/s41589-018-0108-2</a>.","ista":"Zhang W, Herde M, Mitchell J, Whitfield J, Wulff A, Vongsouthi V, Sanchez-Romero I, Gulakova P, Minge D, Breithausen B, Schoch S, Janovjak HL, Jackson C, Henneberger C. 2018. Monitoring hippocampal glycine with the computationally designed optical sensor GlyFS. Nature Chemical Biology. 14(9), 861–869.","short":"W. Zhang, M. Herde, J. Mitchell, J. Whitfield, A. Wulff, V. Vongsouthi, I. Sanchez-Romero, P. Gulakova, D. Minge, B. Breithausen, S. Schoch, H.L. Janovjak, C. Jackson, C. Henneberger, Nature Chemical Biology 14 (2018) 861–869.","apa":"Zhang, W., Herde, M., Mitchell, J., Whitfield, J., Wulff, A., Vongsouthi, V., … Henneberger, C. (2018). Monitoring hippocampal glycine with the computationally designed optical sensor GlyFS. <i>Nature Chemical Biology</i>. Nature Publishing Group. <a href=\"https://doi.org/10.1038/s41589-018-0108-2\">https://doi.org/10.1038/s41589-018-0108-2</a>"},"pmid":1,"type":"journal_article","scopus_import":"1","month":"07","date_published":"2018-07-30T00:00:00Z","article_type":"original","publist_id":"7786","doi":"10.1038/s41589-018-0108-2","oa_version":"Submitted Version"},{"isi":1,"day":"01","department":[{"_id":"HaJa"}],"quality_controlled":"1","publisher":"Elsevier","page":"8 - 14","status":"public","date_created":"2018-12-11T11:49:45Z","abstract":[{"text":"The optogenetic revolution enabled spatially-precise and temporally-precise control over protein function, signaling pathway activation, and animal behavior with tremendous success in the dissection of signaling networks and neural circuits. Very recently, optogenetic methods have been paired with optical reporters in novel drug screening platforms. In these all-optical platforms, light remotely activated ion channels and kinases thereby obviating the use of electrophysiology or reagents. Consequences were remarkable operational simplicity, throughput, and cost-effectiveness that culminated in the identification of new drug candidates. These blueprints for all-optical assays also revealed potential pitfalls and inspire all-optical variants of other screens, such as those that aim at better understanding dynamic drug action or orphan protein function.","lang":"eng"}],"_id":"1026","user_id":"c635000d-4b10-11ee-a964-aac5a93f6ac1","publication_status":"published","author":[{"full_name":"Agus, Viviana","last_name":"Agus","first_name":"Viviana"},{"orcid":"0000-0002-8023-9315","last_name":"Janovjak","first_name":"Harald L","id":"33BA6C30-F248-11E8-B48F-1D18A9856A87","full_name":"Janovjak, Harald L"}],"external_id":{"isi":["000418313200003"]},"project":[{"grant_number":"RGY0084/2012","_id":"255BFFFA-B435-11E9-9278-68D0E5697425","name":"In situ real-time imaging of neurotransmitter signaling using designer optical sensors (HFSP Young Investigator)"},{"grant_number":"303564","call_identifier":"FP7","name":"Microbial Ion Channels for Synthetic Neurobiology","_id":"25548C20-B435-11E9-9278-68D0E5697425"},{"name":"Molecular Drug Targets","_id":"255A6082-B435-11E9-9278-68D0E5697425","grant_number":"W1232-B24","call_identifier":"FWF"}],"volume":48,"publication_identifier":{"issn":["09581669"]},"acknowledgement":"This work was supported by grants of the European Union Seventh Framework Programme (CIG-303564), the Human Frontier Science Program (RGY0084_2012), and the Austrian Science Fund FWF (W1232 MolecularDrugTargets).","intvolume":"        48","title":"Optogenetic methods in drug screening: Technologies and applications","article_processing_charge":"No","publication":"Current Opinion in Biotechnology","language":[{"iso":"eng"}],"year":"2017","citation":{"ama":"Agus V, Janovjak HL. Optogenetic methods in drug screening: Technologies and applications. <i>Current Opinion in Biotechnology</i>. 2017;48:8-14. doi:<a href=\"https://doi.org/10.1016/j.copbio.2017.02.006\">10.1016/j.copbio.2017.02.006</a>","mla":"Agus, Viviana, and Harald L. Janovjak. “Optogenetic Methods in Drug Screening: Technologies and Applications.” <i>Current Opinion in Biotechnology</i>, vol. 48, Elsevier, 2017, pp. 8–14, doi:<a href=\"https://doi.org/10.1016/j.copbio.2017.02.006\">10.1016/j.copbio.2017.02.006</a>.","ieee":"V. Agus and H. L. Janovjak, “Optogenetic methods in drug screening: Technologies and applications,” <i>Current Opinion in Biotechnology</i>, vol. 48. Elsevier, pp. 8–14, 2017.","short":"V. Agus, H.L. Janovjak, Current Opinion in Biotechnology 48 (2017) 8–14.","apa":"Agus, V., &#38; Janovjak, H. L. (2017). Optogenetic methods in drug screening: Technologies and applications. <i>Current Opinion in Biotechnology</i>. Elsevier. <a href=\"https://doi.org/10.1016/j.copbio.2017.02.006\">https://doi.org/10.1016/j.copbio.2017.02.006</a>","chicago":"Agus, Viviana, and Harald L Janovjak. “Optogenetic Methods in Drug Screening: Technologies and Applications.” <i>Current Opinion in Biotechnology</i>. Elsevier, 2017. <a href=\"https://doi.org/10.1016/j.copbio.2017.02.006\">https://doi.org/10.1016/j.copbio.2017.02.006</a>.","ista":"Agus V, Janovjak HL. 2017. Optogenetic methods in drug screening: Technologies and applications. Current Opinion in Biotechnology. 48, 8–14."},"date_updated":"2023-09-22T09:26:06Z","type":"journal_article","date_published":"2017-12-01T00:00:00Z","month":"12","scopus_import":"1","doi":"10.1016/j.copbio.2017.02.006","publist_id":"6365","article_type":"original","ec_funded":1,"oa_version":"None"},{"title":"Ancestral protein reconstruction and circular permutation for improving the stability and dynamic range of FRET sensors","intvolume":"      1596","publication":"Synthetic Protein Switches","language":[{"iso":"eng"}],"year":"2017","type":"book_chapter","citation":{"ama":"Clifton B, Whitfield J, Sanchez-Romero I, et al. Ancestral protein reconstruction and circular permutation for improving the stability and dynamic range of FRET sensors. In: Stein V, ed. <i>Synthetic Protein Switches</i>. Vol 1596. Synthetic Protein Switches. Springer; 2017:71-87. doi:<a href=\"https://doi.org/10.1007/978-1-4939-6940-1_5\">10.1007/978-1-4939-6940-1_5</a>","mla":"Clifton, Ben, et al. “Ancestral Protein Reconstruction and Circular Permutation for Improving the Stability and Dynamic Range of FRET Sensors.” <i>Synthetic Protein Switches</i>, edited by Viktor Stein, vol. 1596, Springer, 2017, pp. 71–87, doi:<a href=\"https://doi.org/10.1007/978-1-4939-6940-1_5\">10.1007/978-1-4939-6940-1_5</a>.","ieee":"B. Clifton <i>et al.</i>, “Ancestral protein reconstruction and circular permutation for improving the stability and dynamic range of FRET sensors,” in <i>Synthetic Protein Switches</i>, vol. 1596, V. Stein, Ed. Springer, 2017, pp. 71–87.","short":"B. Clifton, J. Whitfield, I. Sanchez-Romero, M. Herde, C. Henneberger, H.L. Janovjak, C. Jackson, in:, V. Stein (Ed.), Synthetic Protein Switches, Springer, 2017, pp. 71–87.","apa":"Clifton, B., Whitfield, J., Sanchez-Romero, I., Herde, M., Henneberger, C., Janovjak, H. L., &#38; Jackson, C. (2017). Ancestral protein reconstruction and circular permutation for improving the stability and dynamic range of FRET sensors. In V. Stein (Ed.), <i>Synthetic Protein Switches</i> (Vol. 1596, pp. 71–87). Springer. <a href=\"https://doi.org/10.1007/978-1-4939-6940-1_5\">https://doi.org/10.1007/978-1-4939-6940-1_5</a>","chicago":"Clifton, Ben, Jason Whitfield, Inmaculada Sanchez-Romero, Michel Herde, Christian Henneberger, Harald L Janovjak, and Colin Jackson. “Ancestral Protein Reconstruction and Circular Permutation for Improving the Stability and Dynamic Range of FRET Sensors.” In <i>Synthetic Protein Switches</i>, edited by Viktor Stein, 1596:71–87. Synthetic Protein Switches. Springer, 2017. <a href=\"https://doi.org/10.1007/978-1-4939-6940-1_5\">https://doi.org/10.1007/978-1-4939-6940-1_5</a>.","ista":"Clifton B, Whitfield J, Sanchez-Romero I, Herde M, Henneberger C, Janovjak HL, Jackson C. 2017.Ancestral protein reconstruction and circular permutation for improving the stability and dynamic range of FRET sensors. In: Synthetic Protein Switches. Methods in Molecular Biology, vol. 1596, 71–87."},"date_updated":"2021-01-12T08:22:13Z","date_published":"2017-03-15T00:00:00Z","month":"03","scopus_import":1,"doi":"10.1007/978-1-4939-6940-1_5","publist_id":"6451","editor":[{"first_name":"Viktor","last_name":"Stein","full_name":"Stein, Viktor"}],"alternative_title":["Methods in Molecular Biology"],"oa_version":"None","day":"15","series_title":"Synthetic Protein Switches","department":[{"_id":"HaJa"}],"quality_controlled":"1","publisher":"Springer","page":"71 - 87","date_created":"2018-12-11T11:49:24Z","status":"public","abstract":[{"text":"Small molecule biosensors based on Forster resonance energy transfer (FRET) enable small molecule signaling to be monitored with high spatial and temporal resolution in complex cellular environments. FRET sensors can be constructed by fusing a pair of fluorescent proteins to a suitable recognition domain, such as a member of the solute-binding protein (SBP) superfamily. However, naturally occurring SBPs may be unsuitable for incorporation into FRET sensors due to their low thermostability, which may preclude imaging under physiological conditions, or because the positions of their N- and C-termini may be suboptimal for fusion of fluorescent proteins, which may limit the dynamic range of the resulting sensors. Here, we show how these problems can be overcome using ancestral protein reconstruction and circular permutation. Ancestral protein reconstruction, used as a protein engineering strategy, leverages phylogenetic information to improve the thermostability of proteins, while circular permutation enables the termini of an SBP to be repositioned to maximize the dynamic range of the resulting FRET sensor. We also provide a protocol for cloning the engineered SBPs into FRET sensor constructs using Golden Gate assembly and discuss considerations for in situ characterization of the FRET sensors.","lang":"eng"}],"_id":"957","publication_status":"published","user_id":"4435EBFC-F248-11E8-B48F-1D18A9856A87","author":[{"last_name":"Clifton","first_name":"Ben","full_name":"Clifton, Ben"},{"last_name":"Whitfield","first_name":"Jason","full_name":"Whitfield, Jason"},{"full_name":"Sanchez Romero, Inmaculada","last_name":"Sanchez Romero","first_name":"Inmaculada","id":"3D9C5D30-F248-11E8-B48F-1D18A9856A87"},{"first_name":"Michel","last_name":"Herde","full_name":"Herde, Michel"},{"full_name":"Henneberger, Christian","first_name":"Christian","last_name":"Henneberger"},{"full_name":"Janovjak, Harald L","id":"33BA6C30-F248-11E8-B48F-1D18A9856A87","first_name":"Harald L","orcid":"0000-0002-8023-9315","last_name":"Janovjak"},{"full_name":"Jackson, Colin","first_name":"Colin","last_name":"Jackson"}],"project":[{"_id":"255BFFFA-B435-11E9-9278-68D0E5697425","name":"In situ real-time imaging of neurotransmitter signaling using designer optical sensors (HFSP Young Investigator)","grant_number":"RGY0084/2012"}],"publication_identifier":{"issn":["10643745"]},"volume":1596},{"author":[{"id":"33BA6C30-F248-11E8-B48F-1D18A9856A87","last_name":"Janovjak","orcid":"0000-0002-8023-9315","first_name":"Harald L","full_name":"Janovjak, Harald L"}],"publist_id":"5756","doi":"10.1016/j.str.2016.01.002","ec_funded":1,"oa_version":"None","project":[{"_id":"255BFFFA-B435-11E9-9278-68D0E5697425","name":"In situ real-time imaging of neurotransmitter signaling using designer optical sensors (HFSP Young Investigator)","grant_number":"RGY0084/2012"},{"name":"Microbial Ion Channels for Synthetic Neurobiology","_id":"25548C20-B435-11E9-9278-68D0E5697425","grant_number":"303564","call_identifier":"FP7"},{"_id":"255A6082-B435-11E9-9278-68D0E5697425","name":"Molecular Drug Targets","call_identifier":"FWF","grant_number":"W1232-B24"}],"volume":24,"date_updated":"2021-01-12T06:50:46Z","citation":{"apa":"Janovjak, H. L. (2016). Light at the end of the protein: Crystal structure of a C-terminal light-sensing domain. <i>Structure</i>. Cell Press. <a href=\"https://doi.org/10.1016/j.str.2016.01.002\">https://doi.org/10.1016/j.str.2016.01.002</a>","short":"H.L. Janovjak, Structure 24 (2016) 213–215.","ista":"Janovjak HL. 2016. Light at the end of the protein: Crystal structure of a C-terminal light-sensing domain. Structure. 24(2), 213–215.","chicago":"Janovjak, Harald L. “Light at the End of the Protein: Crystal Structure of a C-Terminal Light-Sensing Domain.” <i>Structure</i>. Cell Press, 2016. <a href=\"https://doi.org/10.1016/j.str.2016.01.002\">https://doi.org/10.1016/j.str.2016.01.002</a>.","ama":"Janovjak HL. Light at the end of the protein: Crystal structure of a C-terminal light-sensing domain. <i>Structure</i>. 2016;24(2):213-215. doi:<a href=\"https://doi.org/10.1016/j.str.2016.01.002\">10.1016/j.str.2016.01.002</a>","ieee":"H. L. Janovjak, “Light at the end of the protein: Crystal structure of a C-terminal light-sensing domain,” <i>Structure</i>, vol. 24, no. 2. Cell Press, pp. 213–215, 2016.","mla":"Janovjak, Harald L. “Light at the End of the Protein: Crystal Structure of a C-Terminal Light-Sensing Domain.” <i>Structure</i>, vol. 24, no. 2, Cell Press, 2016, pp. 213–15, doi:<a href=\"https://doi.org/10.1016/j.str.2016.01.002\">10.1016/j.str.2016.01.002</a>."},"type":"journal_article","_id":"1440","date_published":"2016-02-02T00:00:00Z","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","month":"02","publication_status":"published","scopus_import":1,"page":"213 - 215","publisher":"Cell Press","publication":"Structure","status":"public","date_created":"2018-12-11T11:52:02Z","year":"2016","language":[{"iso":"eng"}],"day":"02","acknowledgement":"The author thanks Banerjee et al. (2016) for providing coordinates prior to public release and apologizes to colleagues whose work was not cited or discussed due to the limited space available. The author is supported by grants from EU FP7 (CIG-303564), HFSP (RGY0084_2012), and FWF (W1232).","department":[{"_id":"HaJa"}],"intvolume":"        24","quality_controlled":"1","title":"Light at the end of the protein: Crystal structure of a C-terminal light-sensing domain","issue":"2"},{"intvolume":"        24","main_file_link":[{"open_access":"1","url":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4570536/"}],"title":"Construction of a robust and sensitive arginine biosensor through ancestral protein reconstruction","year":"2015","language":[{"iso":"eng"}],"publication":"Protein Science","scopus_import":1,"month":"09","date_published":"2015-09-01T00:00:00Z","date_updated":"2021-01-12T06:52:00Z","citation":{"ama":"Whitfield J, Zhang W, Herde M, et al. Construction of a robust and sensitive arginine biosensor through ancestral protein reconstruction. <i>Protein Science</i>. 2015;24(9):1412-1422. doi:<a href=\"https://doi.org/10.1002/pro.2721\">10.1002/pro.2721</a>","ieee":"J. Whitfield <i>et al.</i>, “Construction of a robust and sensitive arginine biosensor through ancestral protein reconstruction,” <i>Protein Science</i>, vol. 24, no. 9. Wiley, pp. 1412–1422, 2015.","mla":"Whitfield, Jason, et al. “Construction of a Robust and Sensitive Arginine Biosensor through Ancestral Protein Reconstruction.” <i>Protein Science</i>, vol. 24, no. 9, Wiley, 2015, pp. 1412–22, doi:<a href=\"https://doi.org/10.1002/pro.2721\">10.1002/pro.2721</a>.","apa":"Whitfield, J., Zhang, W., Herde, M., Clifton, B., Radziejewski, J., Janovjak, H. L., … Jackson, C. (2015). Construction of a robust and sensitive arginine biosensor through ancestral protein reconstruction. <i>Protein Science</i>. Wiley. <a href=\"https://doi.org/10.1002/pro.2721\">https://doi.org/10.1002/pro.2721</a>","short":"J. Whitfield, W. Zhang, M. Herde, B. Clifton, J. Radziejewski, H.L. Janovjak, C. Henneberger, C. Jackson, Protein Science 24 (2015) 1412–1422.","ista":"Whitfield J, Zhang W, Herde M, Clifton B, Radziejewski J, Janovjak HL, Henneberger C, Jackson C. 2015. Construction of a robust and sensitive arginine biosensor through ancestral protein reconstruction. Protein Science. 24(9), 1412–1422.","chicago":"Whitfield, Jason, William Zhang, Michel Herde, Ben Clifton, Johanna Radziejewski, Harald L Janovjak, Christian Henneberger, and Colin Jackson. “Construction of a Robust and Sensitive Arginine Biosensor through Ancestral Protein Reconstruction.” <i>Protein Science</i>. Wiley, 2015. <a href=\"https://doi.org/10.1002/pro.2721\">https://doi.org/10.1002/pro.2721</a>."},"type":"journal_article","pmid":1,"oa_version":"Submitted Version","publist_id":"5555","doi":"10.1002/pro.2721","issue":"9","oa":1,"quality_controlled":"1","department":[{"_id":"HaJa"}],"day":"01","status":"public","date_created":"2018-12-11T11:53:01Z","publisher":"Wiley","page":"1412 - 1422","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","publication_status":"published","_id":"1611","abstract":[{"lang":"eng","text":"Biosensors for signaling molecules allow the study of physiological processes by bringing together the fields of protein engineering, fluorescence imaging, and cell biology. Construction of genetically encoded biosensors generally relies on the availability of a binding &quot;core&quot; that is both specific and stable, which can then be combined with fluorescent molecules to create a sensor. However, binding proteins with the desired properties are often not available in nature and substantial improvement to sensors can be required, particularly with regard to their durability. Ancestral protein reconstruction is a powerful protein-engineering tool able to generate highly stable and functional proteins. In this work, we sought to establish the utility of ancestral protein reconstruction to biosensor development, beginning with the construction of an l-arginine biosensor. l-arginine, as the immediate precursor to nitric oxide, is an important molecule in many physiological contexts including brain function. Using a combination of ancestral reconstruction and circular permutation, we constructed a Förster resonance energy transfer (FRET) biosensor for l-arginine (cpFLIPR). cpFLIPR displays high sensitivity and specificity, with a Kd of ∼14 μM and a maximal dynamic range of 35%. Importantly, cpFLIPR was highly robust, enabling accurate l-arginine measurement at physiological temperatures. We established that cpFLIPR is compatible with two-photon excitation fluorescence microscopy and report l-arginine concentrations in brain tissue."}],"volume":24,"project":[{"name":"In situ real-time imaging of neurotransmitter signaling using designer optical sensors (HFSP Young Investigator)","_id":"255BFFFA-B435-11E9-9278-68D0E5697425","grant_number":"RGY0084/2012"}],"author":[{"full_name":"Whitfield, Jason","last_name":"Whitfield","first_name":"Jason"},{"first_name":"William","last_name":"Zhang","full_name":"Zhang, William"},{"full_name":"Herde, Michel","last_name":"Herde","first_name":"Michel"},{"full_name":"Clifton, Ben","last_name":"Clifton","first_name":"Ben"},{"full_name":"Radziejewski, Johanna","last_name":"Radziejewski","first_name":"Johanna"},{"full_name":"Janovjak, Harald L","id":"33BA6C30-F248-11E8-B48F-1D18A9856A87","first_name":"Harald L","last_name":"Janovjak","orcid":"0000-0002-8023-9315"},{"full_name":"Henneberger, Christian","first_name":"Christian","last_name":"Henneberger"},{"last_name":"Jackson","first_name":"Colin","full_name":"Jackson, Colin"}],"external_id":{"pmid":["26061224"]}},{"day":"12","department":[{"_id":"HaJa"},{"_id":"LifeSc"}],"quality_controlled":"1","oa":1,"issue":"12","file":[{"date_created":"2018-12-12T10:10:51Z","checksum":"e9fb251dfcb7cd209b83f17867e61321","access_level":"open_access","file_name":"IST-2017-837-v1+1_ingles-prieto.pdf","file_id":"4842","relation":"main_file","content_type":"application/pdf","file_size":1308364,"creator":"system","date_updated":"2020-07-14T12:45:12Z"}],"publisher":"Nature Publishing Group","page":"952 - 954","date_created":"2018-12-11T11:53:25Z","status":"public","abstract":[{"text":"High-throughput live-cell screens are intricate elements of systems biology studies and drug discovery pipelines. Here, we demonstrate an optogenetics-assisted method that avoids the need for chemical activators and reporters, reduces the number of operational steps and increases information content in a cell-based small-molecule screen against human protein kinases, including an orphan receptor tyrosine kinase. This blueprint for all-optical screening can be adapted to many drug targets and cellular processes.","lang":"eng"}],"_id":"1678","publication_status":"published","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","author":[{"full_name":"Inglés Prieto, Álvaro","id":"2A9DB292-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0002-5409-8571","last_name":"Inglés Prieto","first_name":"Álvaro"},{"id":"3FEE232A-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0002-7218-7738","last_name":"Gschaider-Reichhart","first_name":"Eva","full_name":"Gschaider-Reichhart, Eva"},{"last_name":"Muellner","first_name":"Markus","full_name":"Muellner, Markus"},{"full_name":"Nowak, Matthias","id":"30845DAA-F248-11E8-B48F-1D18A9856A87","last_name":"Nowak","first_name":"Matthias"},{"full_name":"Nijman, Sebastian","first_name":"Sebastian","last_name":"Nijman"},{"full_name":"Grusch, Michael","last_name":"Grusch","first_name":"Michael"},{"id":"33BA6C30-F248-11E8-B48F-1D18A9856A87","first_name":"Harald L","last_name":"Janovjak","orcid":"0000-0002-8023-9315","full_name":"Janovjak, Harald L"}],"project":[{"_id":"25548C20-B435-11E9-9278-68D0E5697425","name":"Microbial Ion Channels for Synthetic Neurobiology","call_identifier":"FP7","grant_number":"303564"},{"grant_number":"RGY0084/2012","name":"In situ real-time imaging of neurotransmitter signaling using designer optical sensors (HFSP Young Investigator)","_id":"255BFFFA-B435-11E9-9278-68D0E5697425"},{"_id":"255A6082-B435-11E9-9278-68D0E5697425","name":"Molecular Drug Targets","call_identifier":"FWF","grant_number":"W1232-B24"}],"volume":11,"pubrep_id":"837","acknowledgement":"This work was supported by grants from the European Union Seventh Framework Programme (CIG-303564 to H.J. and ERC-StG-311166 to S.M.B.N.), the Human Frontier Science Program (RGY0084_2012 to H.J.) and the Herzfelder Foundation (to M.G.). A.I.-P. was supported by a Ramon Areces fellowship, and E.R. by the graduate program MolecularDrugTargets (Austrian Science Fund (FWF): W 1232) and a FemTech fellowship (3580812 Austrian Research Promotion Agency).","title":"Light-assisted small-molecule screening against protein kinases","intvolume":"        11","publication":"Nature Chemical Biology","language":[{"iso":"eng"}],"year":"2015","type":"journal_article","has_accepted_license":"1","date_updated":"2023-09-07T12:49:09Z","citation":{"ista":"Inglés Prieto Á, Gschaider-Reichhart E, Muellner M, Nowak M, Nijman S, Grusch M, Janovjak HL. 2015. Light-assisted small-molecule screening against protein kinases. Nature Chemical Biology. 11(12), 952–954.","chicago":"Inglés Prieto, Álvaro, Eva Gschaider-Reichhart, Markus Muellner, Matthias Nowak, Sebastian Nijman, Michael Grusch, and Harald L Janovjak. “Light-Assisted Small-Molecule Screening against Protein Kinases.” <i>Nature Chemical Biology</i>. Nature Publishing Group, 2015. <a href=\"https://doi.org/10.1038/nchembio.1933\">https://doi.org/10.1038/nchembio.1933</a>.","apa":"Inglés Prieto, Á., Gschaider-Reichhart, E., Muellner, M., Nowak, M., Nijman, S., Grusch, M., &#38; Janovjak, H. L. (2015). Light-assisted small-molecule screening against protein kinases. <i>Nature Chemical Biology</i>. Nature Publishing Group. <a href=\"https://doi.org/10.1038/nchembio.1933\">https://doi.org/10.1038/nchembio.1933</a>","short":"Á. Inglés Prieto, E. Gschaider-Reichhart, M. Muellner, M. Nowak, S. Nijman, M. Grusch, H.L. Janovjak, Nature Chemical Biology 11 (2015) 952–954.","ieee":"Á. Inglés Prieto <i>et al.</i>, “Light-assisted small-molecule screening against protein kinases,” <i>Nature Chemical Biology</i>, vol. 11, no. 12. Nature Publishing Group, pp. 952–954, 2015.","mla":"Inglés Prieto, Álvaro, et al. “Light-Assisted Small-Molecule Screening against Protein Kinases.” <i>Nature Chemical Biology</i>, vol. 11, no. 12, Nature Publishing Group, 2015, pp. 952–54, doi:<a href=\"https://doi.org/10.1038/nchembio.1933\">10.1038/nchembio.1933</a>.","ama":"Inglés Prieto Á, Gschaider-Reichhart E, Muellner M, et al. Light-assisted small-molecule screening against protein kinases. <i>Nature Chemical Biology</i>. 2015;11(12):952-954. doi:<a href=\"https://doi.org/10.1038/nchembio.1933\">10.1038/nchembio.1933</a>"},"ddc":["571"],"date_published":"2015-10-12T00:00:00Z","month":"10","scopus_import":1,"doi":"10.1038/nchembio.1933","publist_id":"5471","related_material":{"record":[{"relation":"dissertation_contains","id":"418","status":"public"}]},"ec_funded":1,"oa_version":"Submitted Version","file_date_updated":"2020-07-14T12:45:12Z"},{"day":"01","department":[{"_id":"HaJa"}],"quality_controlled":"1","issue":"4","page":"518 - 525","publisher":"Wiley","status":"public","date_created":"2018-12-11T11:54:26Z","abstract":[{"lang":"eng","text":"Cultured mammalian cells essential are model systems in basic biology research, production platforms of proteins for medical use, and testbeds in synthetic biology. Flavin cofactors, in particular flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), are critical for cellular redox reactions and sense light in naturally occurring photoreceptors and optogenetic tools. Here, we quantified flavin contents of commonly used mammalian cell lines. We first compared three procedures for extraction of free and noncovalently protein-bound flavins and verified extraction using fluorescence spectroscopy. For separation, two CE methods with different BGEs were established, and detection was performed by LED-induced fluorescence with limit of detections (LODs 0.5-3.8 nM). We found that riboflavin (RF), FMN, and FAD contents varied significantly between cell lines. RF (3.1-14 amol/cell) and FAD (2.2-17.0 amol/cell) were the predominant flavins, while FMN (0.46-3.4 amol/cell) was found at markedly lower levels. Observed flavin contents agree with those previously extracted from mammalian tissues, yet reduced forms of RF were detected that were not described previously. Quantification of flavins in mammalian cell lines will allow a better understanding of cellular redox reactions and optogenetic tools."}],"_id":"1867","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","publication_status":"published","author":[{"first_name":"Jens","last_name":"Hühner","full_name":"Hühner, Jens"},{"first_name":"Álvaro","orcid":"0000-0002-5409-8571","last_name":"Inglés Prieto","id":"2A9DB292-F248-11E8-B48F-1D18A9856A87","full_name":"Inglés Prieto, Álvaro"},{"full_name":"Neusüß, Christian","first_name":"Christian","last_name":"Neusüß"},{"last_name":"Lämmerhofer","first_name":"Michael","full_name":"Lämmerhofer, Michael"},{"full_name":"Janovjak, Harald L","first_name":"Harald L","orcid":"0000-0002-8023-9315","last_name":"Janovjak","id":"33BA6C30-F248-11E8-B48F-1D18A9856A87"}],"project":[{"call_identifier":"FP7","grant_number":"303564","_id":"25548C20-B435-11E9-9278-68D0E5697425","name":"Microbial Ion Channels for Synthetic Neurobiology"},{"_id":"255BFFFA-B435-11E9-9278-68D0E5697425","name":"In situ real-time imaging of neurotransmitter signaling using designer optical sensors (HFSP Young Investigator)","grant_number":"RGY0084/2012"}],"volume":36,"pubrep_id":"836","intvolume":"        36","title":"Quantification of riboflavin, flavin mononucleotide, and flavin adenine dinucleotide in mammalian model cells by CE with LED-induced fluorescence detection","publication":"Electrophoresis","language":[{"iso":"eng"}],"year":"2015","date_updated":"2021-01-12T06:53:43Z","citation":{"short":"J. Hühner, Á. Inglés Prieto, C. Neusüß, M. Lämmerhofer, H.L. Janovjak, Electrophoresis 36 (2015) 518–525.","apa":"Hühner, J., Inglés Prieto, Á., Neusüß, C., Lämmerhofer, M., &#38; Janovjak, H. L. (2015). Quantification of riboflavin, flavin mononucleotide, and flavin adenine dinucleotide in mammalian model cells by CE with LED-induced fluorescence detection. <i>Electrophoresis</i>. Wiley. <a href=\"https://doi.org/10.1002/elps.201400451\">https://doi.org/10.1002/elps.201400451</a>","chicago":"Hühner, Jens, Álvaro Inglés Prieto, Christian Neusüß, Michael Lämmerhofer, and Harald L Janovjak. “Quantification of Riboflavin, Flavin Mononucleotide, and Flavin Adenine Dinucleotide in Mammalian Model Cells by CE with LED-Induced Fluorescence Detection.” <i>Electrophoresis</i>. Wiley, 2015. <a href=\"https://doi.org/10.1002/elps.201400451\">https://doi.org/10.1002/elps.201400451</a>.","ista":"Hühner J, Inglés Prieto Á, Neusüß C, Lämmerhofer M, Janovjak HL. 2015. Quantification of riboflavin, flavin mononucleotide, and flavin adenine dinucleotide in mammalian model cells by CE with LED-induced fluorescence detection. Electrophoresis. 36(4), 518–525.","ama":"Hühner J, Inglés Prieto Á, Neusüß C, Lämmerhofer M, Janovjak HL. Quantification of riboflavin, flavin mononucleotide, and flavin adenine dinucleotide in mammalian model cells by CE with LED-induced fluorescence detection. <i>Electrophoresis</i>. 2015;36(4):518-525. doi:<a href=\"https://doi.org/10.1002/elps.201400451\">10.1002/elps.201400451</a>","mla":"Hühner, Jens, et al. “Quantification of Riboflavin, Flavin Mononucleotide, and Flavin Adenine Dinucleotide in Mammalian Model Cells by CE with LED-Induced Fluorescence Detection.” <i>Electrophoresis</i>, vol. 36, no. 4, Wiley, 2015, pp. 518–25, doi:<a href=\"https://doi.org/10.1002/elps.201400451\">10.1002/elps.201400451</a>.","ieee":"J. Hühner, Á. Inglés Prieto, C. Neusüß, M. Lämmerhofer, and H. L. Janovjak, “Quantification of riboflavin, flavin mononucleotide, and flavin adenine dinucleotide in mammalian model cells by CE with LED-induced fluorescence detection,” <i>Electrophoresis</i>, vol. 36, no. 4. Wiley, pp. 518–525, 2015."},"type":"journal_article","month":"02","date_published":"2015-02-01T00:00:00Z","scopus_import":1,"doi":"10.1002/elps.201400451","publist_id":"5230","ec_funded":1,"oa_version":"None"},{"date_created":"2018-12-11T11:59:57Z","status":"public","publisher":"Springer","page":"417 - 435","file":[{"file_id":"4952","relation":"main_file","creator":"system","date_updated":"2020-07-14T12:45:51Z","content_type":"application/pdf","file_size":336734,"date_created":"2018-12-12T10:12:34Z","checksum":"1701f0d989f27ddac471b19a894ec0d1","access_level":"open_access","file_name":"IST-2017-834-v1+1_szobota.pdf"}],"quality_controlled":"1","oa":1,"day":"22","department":[{"_id":"HaJa"}],"project":[{"grant_number":"RGY0084/2012","name":"In situ real-time imaging of neurotransmitter signaling using designer optical sensors (HFSP Young Investigator)","_id":"255BFFFA-B435-11E9-9278-68D0E5697425"},{"name":"Microbial Ion Channels for Synthetic Neurobiology","_id":"25548C20-B435-11E9-9278-68D0E5697425","grant_number":"303564","call_identifier":"FP7"}],"volume":998,"author":[{"last_name":"Szobota","first_name":"Stephanie","full_name":"Szobota, Stephanie"},{"id":"3EEDE19A-F248-11E8-B48F-1D18A9856A87","last_name":"Mckenzie","first_name":"Catherine","full_name":"Mckenzie, Catherine"},{"first_name":"Harald L","last_name":"Janovjak","orcid":"0000-0002-8023-9315","id":"33BA6C30-F248-11E8-B48F-1D18A9856A87","full_name":"Janovjak, Harald L"}],"publication_status":"published","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","abstract":[{"lang":"eng","text":"In the vibrant field of optogenetics, optics and genetic targeting are combined to commandeer cellular functions, such as the neuronal action potential, by optically stimulating light-sensitive ion channels expressed in the cell membrane. One broadly applicable manifestation of this approach are covalently attached photochromic tethered ligands (PTLs) that allow activating ligand-gated ion channels with outstanding spatial and temporal resolution. Here, we describe all steps towards the successful development and application of PTL-gated ion channels in cell lines and primary cells. The basis for these experiments forms a combination of molecular modeling, genetic engineering, cell culture, and electrophysiology. The light-gated glutamate receptor (LiGluR), which consists of the PTL-functionalized GluK2 receptor, serves as a model."}],"_id":"2857","year":"2013","language":[{"iso":"eng"}],"publication":"Methods in Molecular Biology","title":"Optical control of ligand-gated ion channels","intvolume":"       998","pubrep_id":"834","oa_version":"Submitted Version","file_date_updated":"2020-07-14T12:45:51Z","doi":"10.1007/978-1-62703-351-0_32","publist_id":"3932","alternative_title":["MIMB"],"ec_funded":1,"ddc":["570"],"date_published":"2013-02-22T00:00:00Z","month":"02","scopus_import":1,"type":"journal_article","has_accepted_license":"1","citation":{"ama":"Szobota S, Mckenzie C, Janovjak HL. Optical control of ligand-gated ion channels. <i>Methods in Molecular Biology</i>. 2013;998:417-435. doi:<a href=\"https://doi.org/10.1007/978-1-62703-351-0_32\">10.1007/978-1-62703-351-0_32</a>","ieee":"S. Szobota, C. Mckenzie, and H. L. Janovjak, “Optical control of ligand-gated ion channels,” <i>Methods in Molecular Biology</i>, vol. 998. Springer, pp. 417–435, 2013.","mla":"Szobota, Stephanie, et al. “Optical Control of Ligand-Gated Ion Channels.” <i>Methods in Molecular Biology</i>, vol. 998, Springer, 2013, pp. 417–35, doi:<a href=\"https://doi.org/10.1007/978-1-62703-351-0_32\">10.1007/978-1-62703-351-0_32</a>.","apa":"Szobota, S., Mckenzie, C., &#38; Janovjak, H. L. (2013). Optical control of ligand-gated ion channels. <i>Methods in Molecular Biology</i>. Springer. <a href=\"https://doi.org/10.1007/978-1-62703-351-0_32\">https://doi.org/10.1007/978-1-62703-351-0_32</a>","short":"S. Szobota, C. Mckenzie, H.L. Janovjak, Methods in Molecular Biology 998 (2013) 417–435.","ista":"Szobota S, Mckenzie C, Janovjak HL. 2013. Optical control of ligand-gated ion channels. Methods in Molecular Biology. 998, 417–435.","chicago":"Szobota, Stephanie, Catherine Mckenzie, and Harald L Janovjak. “Optical Control of Ligand-Gated Ion Channels.” <i>Methods in Molecular Biology</i>. Springer, 2013. <a href=\"https://doi.org/10.1007/978-1-62703-351-0_32\">https://doi.org/10.1007/978-1-62703-351-0_32</a>."},"date_updated":"2021-01-12T07:00:17Z"}]