Auto-inducing media for uniform isotope labeling of proteins with 15N, 13C and 2H

Auto-inducing media for protein expression offer many advantages like robust reproducibility, high yields of soluble protein and much reduced workload. Here, an auto-inducing medium for uniform isotope labelling of proteins with 15 N, 13 C and/or 2 H in E. coli is presented. So far, auto-inducing me...

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Bibliographic Details
Published in:Journal of biomolecular NMR 2015-06, Vol.62 (2), p.169-177
Main Authors: Guthertz, Nicolas, Klopp, Julia, Winterhalter, Aurélie, Fernández, César, Gossert, Alvar D.
Format: Article
Language:English
Subjects:
Biochemistry
Biological and Medical Physics
Biophysics
Physics
Physics and Astronomy
Spectroscopy/Spectrometry
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Summary:Auto-inducing media for protein expression offer many advantages like robust reproducibility, high yields of soluble protein and much reduced workload. Here, an auto-inducing medium for uniform isotope labelling of proteins with 15 N, 13 C and/or 2 H in E. coli is presented. So far, auto-inducing media have not found widespread application in the NMR field, because of the prohibitively high cost of labeled lactose, which is an essential ingredient of such media. Here, we propose using lactose that is only selectively labeled on the glucose moiety. It can be synthesized from inexpensive and readily available substrates: labeled glucose and unlabeled activated galactose. With this approach, uniformly isotope labeled proteins were expressed in unattended auto-inducing cultures with incorporation of 13 C, 15 N of 96.6 % and 2 H, 15 N of 98.8 %. With the present protocol, the NMR community could profit from the many advantages that auto-inducing media offer.
ISSN:0925-2738
1573-5001
DOI:10.1007/s10858-015-9931-x