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Nucleic Acids Research 2005 33(18):e164; doi:10.1093/nar/gni165
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Published online 27 October 2005

© The Author 2005. Published by Oxford University Press. All rights reserved
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions{at}oxfordjournals.org

Methods Online

Partial 13C isotopic enrichment of nucleoside monophosphates: useful reporters for NMR structural studies

Anita I. Kishore, Michael R. Mayer and James H. Prestegard*

Complex Carbohydrate Research Center, University of Georgia Athens, GA 30602-4712, USA

*To whom correspondence should be addressed. Tel: +1 706 542 6281; Fax: +1 706 542 4412; Email: jpresteg{at}ccrc.uga.edu

Received August 29, 2005. Revised October 3, 2005. Accepted October 3, 2005.

Analysis of the 13C isotopic labeling patterns of nucleoside monophosphates (NMPs) extracted from Escherichia coli grown in a mixture of C-1 and C-2 glucose is presented. By comparing our results to previous observations on amino acids grown in similar media, we have been able to rationalize the labeling pattern based on the well-known biochemistry of nucleotide biosynthesis. Except for a few notable absences of label (C4 in purines and C3' in ribose) and one highly enriched site (C1' in ribose), most carbons are randomly enriched at a low level (an average of 13%). These sparsely labeled NMPs give less complex NMR spectra than their fully isotopically labeled analogs due to the elimination of most 13C–13C scalar couplings. The spectral simplicity is particularly advantageous when working in ordered systems, as illustrated with guanosine diphosphate (GDP) bound to ADP ribosylation factor 1 (ARF1) aligned in a liquid crystalline medium. In this system, the absence of scalar couplings and additional long-range dipolar couplings significantly enhances signal to noise and resolution.


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