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Opdensteinen, P.* ; Sperl, L.E. ; Mohamadi, M. ; Kündgen-Redding, N.* ; Hagn, F. ; Buyel, J.F.*

The transient expression of recombinant proteins in plant cell packs facilitates stable isotope labeling for NMR spectroscopy.

Plant Biotechnol. J. 20, 1928-1939 (2022)
Verlagsversion Forschungsdaten DOI PMC
Open Access Gold
Creative Commons Lizenzvertrag
Nuclear magnetic resonance (NMR) spectroscopy can be used to determine the structure, dynamics and interactions of proteins. However, protein NMR requires stable isotope labeling for signal detection. The cells used for the production of recombinant proteins must therefore be grown in medium containing isotopically labeled substrates. Stable isotope labeling is well established in Escherichia coli, but bacteria are only suitable for the production of simple proteins without post-translational modifications. More complex proteins require eukaryotic production hosts, but their growth can be impaired by labeled media, thus reducing product yields and increasing costs. To address this limitation, we used media supplemented with isotope-labeled substrates to cultivate the tobacco-derived cell line BY-2, which was then cast into plant cell packs (PCPs) for the transient expression of a labeled version of the model protein GB1. Mass spectrometry confirmed the feasibility of isotope labeling with 15 N and 2 H using this approach. The resulting NMR spectrum featured a signal dispersion comparable to recombinant GB1 produced in E. coli. PCPs therefore offer a rapid and cost-efficient alternative for the production of isotope-labeled proteins for NMR analysis, especially suitable for complex proteins that cannot be produced in microbial systems.
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Publikationstyp Artikel: Journalartikel
Dokumenttyp Wissenschaftlicher Artikel
Korrespondenzautor
Schlagwörter Alternative Expression Host ; Defined Cultivation Media ; Isotope Labeling ; Plant Cell Culture ; Structural Analysis
ISSN (print) / ISBN 1467-7644
e-ISSN 1467-7652
Zeitschrift Plant Biotechnology Journal
Quellenangaben Band: 20, Heft: 10, Seiten: 1928-1939 Artikelnummer: , Supplement: ,
Verlag Wiley
Nichtpatentliteratur Publikationen
Begutachtungsstatus Peer reviewed
Institut(e) Institute of Structural Biology (STB)