Abstract
In vivo or whole-cell solid-state NMR is an emerging field which faces tremendous challenges. In most cases, cell biochemistry does not allow the labelling of specific molecules and an in vivo study is thus hindered by the inherent difficulty of identifying, among a formidable number of resonances, those arising from a given molecule. In this work we examined the possibility of studying, by solid-state NMR, the model organism Chlamydomonas reinhardtii fully and non-specifically 13C labelled. The extension of NMR-based dynamic filtering from one-dimensional to two-dimensional experiments enabled an enhanced selectivity which facilitated the assignment of cell constituents. The number of resonances detected with these robust and broadly applicable experiments appears to be surprisingly sparse. Various constituents, notably galactolipids abundant in organelle membranes, carbohydrates from the cell wall, and starch from storage grains could be unambiguously assigned. Moreover, the dominant crystal form of starch could be determined in situ. This work illustrates the feasibility and caveats of using solid-state NMR to study intact non-specifically 13C labelled micro-organisms.
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Acknowledgements
The authors would like to thank Dr. Francesca Zito (CNRS, France) for providing the Chlamydomonas strain and her insights on Chlamydomonas growth and physiology. This work was supported by the Natural Sciences and Engineering Research Council (NSERC) of Canada (Grant 326750-2013 to I.M.) and the Centre National de la Recherche Scientifique (UMR 7099 to D.E.W.). J.-P.B. would like to acknowledge the Groupe de Recherche Axé sur la Structure des Protéines (GRASP) and the NSERC for the award of scholarships. B.G. would like to thank the Canadian Institutes of Health Research Strategic Training initiative in Chemical Biology and the Réseau Aquaculture Québec (RAQ) for the award of scholarships. IM and RT are members of the RAQ.
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Arnold, A.A., Bourgouin, JP., Genard, B. et al. Whole cell solid-state NMR study of Chlamydomonas reinhardtii microalgae. J Biomol NMR 70, 123–131 (2018). https://doi.org/10.1007/s10858-018-0164-7
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DOI: https://doi.org/10.1007/s10858-018-0164-7