caa a22 4500 445325402 CHVBK 20180317142718.0 cr unu---uuuuu 170323e20111201xx s 000 0 eng 10.1007/s10858-011-9570-9 doi (NATIONALLICENCE)springer-10.1007/s10858-011-9570-9 A simple protocol for amino acid type selective isotope labeling in insect cells with improved yields and high reproducibility [Elektronische Daten] [Alvar Gossert, Alexandra Hinniger, Sascha Gutmann, Wolfgang Jahnke, André Strauss, César Fernández] An easy to use and robust approach for amino acid type selective isotope labeling in insect cells is presented. It relies on inexpensive commercial media and can be implemented in laboratories without sophisticated infrastructure. In contrast to previous protocols, where either high protein amounts or high incorporation ratios were obtained, here we achieve both at the same time. By supplementing media with a well considered amount of yeast extract, similar protein amounts as with full media are obtained, without compromising on isotope incorporation. In single and dual amino acid labeling experiments incorporation ratios are consistently ≥90% for all amino acids tested. This enables NMR studies of eukaryotic proteins and their interactions even for proteins with low expression levels. We show applications with human kinases, where protein-ligand interactions are characterized by 2D [15N, 1H]- and [13C, 1H]-HSQC spectra. Springer Science+Business Media B.V., 2011 Isotope labeling nationallicence Amino acid selective nationallicence Insect cells nationallicence Baculovirus nationallicence Kinase nationallicence NMR nationallicence Gossert Alvar Novartis Institutes for BioMedical Research, Novartis AG, 4002, Basel, Switzerland aut Hinniger Alexandra Novartis Institutes for BioMedical Research, Novartis AG, 4002, Basel, Switzerland aut Gutmann Sascha Novartis Institutes for BioMedical Research, Novartis AG, 4002, Basel, Switzerland aut Jahnke Wolfgang Novartis Institutes for BioMedical Research, Novartis AG, 4002, Basel, Switzerland aut Strauss André Novartis Institutes for BioMedical Research, Novartis AG, 4002, Basel, Switzerland aut Fernández César Novartis Institutes for BioMedical Research, Novartis AG, 4002, Basel, Switzerland aut Journal of Biomolecular NMR Springer Netherlands 51/4(2011-12-01), 449-456 0925-2738 51:4<449 2011 51 10858 https://doi.org/10.1007/s10858-011-9570-9 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s10858-011-9570-9 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Gossert Alvar Novartis Institutes for BioMedical Research, Novartis AG, 4002, Basel, Switzerland aut NATIONALLICENCE 700 1- Hinniger Alexandra Novartis Institutes for BioMedical Research, Novartis AG, 4002, Basel, Switzerland aut NATIONALLICENCE 700 1- Gutmann Sascha Novartis Institutes for BioMedical Research, Novartis AG, 4002, Basel, Switzerland aut NATIONALLICENCE 700 1- Jahnke Wolfgang Novartis Institutes for BioMedical Research, Novartis AG, 4002, Basel, Switzerland aut NATIONALLICENCE 700 1- Strauss André Novartis Institutes for BioMedical Research, Novartis AG, 4002, Basel, Switzerland aut NATIONALLICENCE 700 1- Fernández César Novartis Institutes for BioMedical Research, Novartis AG, 4002, Basel, Switzerland aut NATIONALLICENCE 773 0- Journal of Biomolecular NMR Springer Netherlands 51/4(2011-12-01), 449-456 0925-2738 51:4<449 2011 51 10858 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer