Site-selective 13C labeling of histidine and tryptophan using ribose
(2017) In Journal of Biomolecular NMR 69(1). p.23-30- Abstract
Experimental studies on protein dynamics at atomic resolution by NMR-spectroscopy in solution require isolated 1H-X spin pairs. This is the default scenario in standard 1H-15N backbone experiments. Side chain dynamic experiments, which allow to study specific local processes like proton-transfer, or tautomerization, require isolated 1H-13C sites which must be produced by site-selective 13C labeling. In the most general way this is achieved by using site-selectively 13C-enriched glucose as the carbon source in bacterial expression systems. Here we systematically investigate the use of site-selectively 13C-enriched ribose as a suitable precursor for... (More)
Experimental studies on protein dynamics at atomic resolution by NMR-spectroscopy in solution require isolated 1H-X spin pairs. This is the default scenario in standard 1H-15N backbone experiments. Side chain dynamic experiments, which allow to study specific local processes like proton-transfer, or tautomerization, require isolated 1H-13C sites which must be produced by site-selective 13C labeling. In the most general way this is achieved by using site-selectively 13C-enriched glucose as the carbon source in bacterial expression systems. Here we systematically investigate the use of site-selectively 13C-enriched ribose as a suitable precursor for 13C labeled histidines and tryptophans. The 13C incorporation in nearly all sites of all 20 amino acids was quantified and compared to glucose based labeling. In general the ribose approach results in more selective labeling. 1-13C ribose exclusively labels His δ2 and Trp δ1 in aromatic side chains and helps to resolve possible overlap problems. The incorporation yield is however only 37% in total and 72% compared to yields of 2-13C glucose. A combined approach of 1-13C ribose and 2-13C glucose maximizes 13C incorporation to 75% in total and 150% compared to 2-13C glucose only. Further histidine positions β, α and CO become significantly labeled at around 50% in total by 3-, 4- or 5-13C ribose. Interestingly backbone CO of Gly, Ala, Cys, Ser, Val, Phe and Tyr are labeled at 40–50% in total with 3-13C ribose, compared to 5% and below for 1-13C and 2-13C glucose. Using ribose instead of glucose as a source for site-selective 13C labeling enables a very selective labeling of certain positions and thereby expanding the toolbox for customized isotope labeling of amino-acids.
(Less)
- author
- Weininger, Ulrich LU
- organization
- publishing date
- 2017-09
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- Aromatic side chain, Isotope labeling, NMR, Protein dynamics, Relaxation
- in
- Journal of Biomolecular NMR
- volume
- 69
- issue
- 1
- pages
- 23 - 30
- publisher
- Springer
- external identifiers
-
- scopus:85028595953
- pmid:28856561
- wos:000412311500003
- ISSN
- 0925-2738
- DOI
- 10.1007/s10858-017-0130-9
- language
- English
- LU publication?
- yes
- id
- ee561a0b-11b8-4c41-96bc-5dec1071f29e
- date added to LUP
- 2017-09-26 14:59:50
- date last changed
- 2024-04-14 18:18:52
@article{ee561a0b-11b8-4c41-96bc-5dec1071f29e,
abstract = {{<p>Experimental studies on protein dynamics at atomic resolution by NMR-spectroscopy in solution require isolated <sup>1</sup>H-X spin pairs. This is the default scenario in standard <sup>1</sup>H-<sup>15</sup>N backbone experiments. Side chain dynamic experiments, which allow to study specific local processes like proton-transfer, or tautomerization, require isolated <sup>1</sup>H-<sup>13</sup>C sites which must be produced by site-selective <sup>13</sup>C labeling. In the most general way this is achieved by using site-selectively <sup>13</sup>C-enriched glucose as the carbon source in bacterial expression systems. Here we systematically investigate the use of site-selectively <sup>13</sup>C-enriched ribose as a suitable precursor for <sup>13</sup>C labeled histidines and tryptophans. The <sup>13</sup>C incorporation in nearly all sites of all 20 amino acids was quantified and compared to glucose based labeling. In general the ribose approach results in more selective labeling. 1-<sup>13</sup>C ribose exclusively labels His δ2 and Trp δ1 in aromatic side chains and helps to resolve possible overlap problems. The incorporation yield is however only 37% in total and 72% compared to yields of 2-<sup>13</sup>C glucose. A combined approach of 1-<sup>13</sup>C ribose and 2-<sup>13</sup>C glucose maximizes <sup>13</sup>C incorporation to 75% in total and 150% compared to 2-<sup>13</sup>C glucose only. Further histidine positions β, α and CO become significantly labeled at around 50% in total by 3-, 4- or 5-<sup>13</sup>C ribose. Interestingly backbone CO of Gly, Ala, Cys, Ser, Val, Phe and Tyr are labeled at 40–50% in total with 3-<sup>13</sup>C ribose, compared to 5% and below for 1-<sup>13</sup>C and 2-<sup>13</sup>C glucose. Using ribose instead of glucose as a source for site-selective <sup>13</sup>C labeling enables a very selective labeling of certain positions and thereby expanding the toolbox for customized isotope labeling of amino-acids.</p>}},
author = {{Weininger, Ulrich}},
issn = {{0925-2738}},
keywords = {{Aromatic side chain; Isotope labeling; NMR; Protein dynamics; Relaxation}},
language = {{eng}},
number = {{1}},
pages = {{23--30}},
publisher = {{Springer}},
series = {{Journal of Biomolecular NMR}},
title = {{Site-selective <sup>13</sup>C labeling of histidine and tryptophan using ribose}},
url = {{http://dx.doi.org/10.1007/s10858-017-0130-9}},
doi = {{10.1007/s10858-017-0130-9}},
volume = {{69}},
year = {{2017}},
}