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Enabling NMR Studies of High Molecular Weight Systems Without the Need for Deuteration: The XL‐ALSOFAST Experiment with Delayed Decoupling
Author(s) -
Rößler Philip,
Mathieu Daniel,
Gossert Alvar D.
Publication year - 2020
Publication title -
angewandte chemie international edition
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.831
H-Index - 550
eISSN - 1521-3773
pISSN - 1433-7851
DOI - 10.1002/anie.202007715
Subject(s) - decoupling (probability) , nuclear magnetic resonance spectroscopy , protonation , chemistry , relaxation (psychology) , chemical physics , biophysics , nuclear magnetic resonance , biological system , physics , stereochemistry , biology , organic chemistry , engineering , ion , control engineering , neuroscience
Current biological research increasingly focusses on large human proteins and their complexes. Such proteins are difficult to study by NMR spectroscopy because they often can only be produced in higher eukaryotic expression systems, where deuteration is hardly feasible. Here, we present the XL‐ALSOFAST‐[ 13 C, 1 H]‐HMQC experiment with much improved sensitivity for fully protonated high molecular weight proteins. For the tested systems ranging from 100 to 240 kDa in size, 3‐fold higher sensitivity was obtained on average for fast relaxing signals compared to current state‐of‐the‐art experiments. In the XL‐ALSOFAST approach, non‐observed magnetisation is optimally exploited and transverse relaxation is minimized by the newly introduced concept of delayed decoupling. The combination of high sensitivity and superior artefact suppression makes it ideal for studying inherently unstable membrane proteins or for analysing therapeutic antibodies at natural 13 C abundance. The XL‐ALSOFAST and delayed decoupling will therefore expand the range of biomolecular systems accessible to NMR spectroscopy.