Site-selective 13C labeling of proteins using erythrose
(2017) In Journal of Biomolecular NMR 67(3). p.191-200- Abstract
NMR-spectroscopy enables unique experimental studies on protein dynamics at atomic resolution. In order to obtain a full atom view on protein dynamics, and to study specific local processes like ring-flips, proton-transfer, or tautomerization, one has to perform studies on amino-acid side chains. A key requirement for these studies is site-selective labeling with 13C and/or 1H, which is achieved in the most general way by using site-selectively 13C-enriched glucose (1- and 2-13C) as the carbon source in bacterial expression systems. Using this strategy, multiple sites in side chains, including aromatics, become site-selectively labeled and suitable for relaxation studies. Here we... (More)
NMR-spectroscopy enables unique experimental studies on protein dynamics at atomic resolution. In order to obtain a full atom view on protein dynamics, and to study specific local processes like ring-flips, proton-transfer, or tautomerization, one has to perform studies on amino-acid side chains. A key requirement for these studies is site-selective labeling with 13C and/or 1H, which is achieved in the most general way by using site-selectively 13C-enriched glucose (1- and 2-13C) as the carbon source in bacterial expression systems. Using this strategy, multiple sites in side chains, including aromatics, become site-selectively labeled and suitable for relaxation studies. Here we systematically investigate the use of site-selectively 13C-enriched erythrose (1-, 2-, 3- and 4-13C) as a suitable precursor for 13C labeled aromatic side chains. We quantify 13C incorporation in nearly all sites in all 20 amino acids and compare the results to glucose based labeling. In general the erythrose approach results in more selective labeling. While there is only a minor gain for phenylalanine and tyrosine side-chains, the 13C incorporation level for tryptophan is at least doubled. Additionally, the Phe ζ and Trp η2 positions become labeled. In the aliphatic side chains, labeling using erythrose yields isolated 13C labels for certain positions, like Ile β and His β, making these sites suitable for dynamics studies. Using erythrose instead of glucose as a source for site-selective 13C labeling enables unique or superior labeling for certain positions and is thereby expanding the toolbox for customized isotope labeling of amino-acid side-chains.
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- author
- Weininger, Ulrich LU
- organization
- publishing date
- 2017-02-28
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- Aromatic side chain, Isotope labeling, Protein dynamics, Relaxation
- in
- Journal of Biomolecular NMR
- volume
- 67
- issue
- 3
- pages
- 191 - 200
- publisher
- Springer
- external identifiers
-
- scopus:85014031823
- pmid:28247186
- wos:000399299600003
- ISSN
- 0925-2738
- DOI
- 10.1007/s10858-017-0096-7
- language
- English
- LU publication?
- yes
- id
- 988a7399-8984-4713-a99f-f6ebfb9750e3
- date added to LUP
- 2017-03-13 10:02:55
- date last changed
- 2024-06-23 13:27:35
@article{988a7399-8984-4713-a99f-f6ebfb9750e3,
abstract = {{<p>NMR-spectroscopy enables unique experimental studies on protein dynamics at atomic resolution. In order to obtain a full atom view on protein dynamics, and to study specific local processes like ring-flips, proton-transfer, or tautomerization, one has to perform studies on amino-acid side chains. A key requirement for these studies is site-selective labeling with <sup>13</sup>C and/or <sup>1</sup>H, which is achieved in the most general way by using site-selectively <sup>13</sup>C-enriched glucose (1- and 2-<sup>13</sup>C) as the carbon source in bacterial expression systems. Using this strategy, multiple sites in side chains, including aromatics, become site-selectively labeled and suitable for relaxation studies. Here we systematically investigate the use of site-selectively <sup>13</sup>C-enriched erythrose (1-, 2-, 3- and 4-<sup>13</sup>C) as a suitable precursor for <sup>13</sup>C labeled aromatic side chains. We quantify <sup>13</sup>C incorporation in nearly all sites in all 20 amino acids and compare the results to glucose based labeling. In general the erythrose approach results in more selective labeling. While there is only a minor gain for phenylalanine and tyrosine side-chains, the <sup>13</sup>C incorporation level for tryptophan is at least doubled. Additionally, the Phe ζ and Trp η2 positions become labeled. In the aliphatic side chains, labeling using erythrose yields isolated <sup>13</sup>C labels for certain positions, like Ile β and His β, making these sites suitable for dynamics studies. Using erythrose instead of glucose as a source for site-selective <sup>13</sup>C labeling enables unique or superior labeling for certain positions and is thereby expanding the toolbox for customized isotope labeling of amino-acid side-chains.</p>}},
author = {{Weininger, Ulrich}},
issn = {{0925-2738}},
keywords = {{Aromatic side chain; Isotope labeling; Protein dynamics; Relaxation}},
language = {{eng}},
month = {{02}},
number = {{3}},
pages = {{191--200}},
publisher = {{Springer}},
series = {{Journal of Biomolecular NMR}},
title = {{Site-selective <sup>13</sup>C labeling of proteins using erythrose}},
url = {{http://dx.doi.org/10.1007/s10858-017-0096-7}},
doi = {{10.1007/s10858-017-0096-7}},
volume = {{67}},
year = {{2017}},
}