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Improved Resolution and Sensitivity of Triple-Resonance NMR Methods for the Structural Analysis of Proteins by Use of a Backbone-Labeling Strategy

Lookup NU author(s): Professor Steve Homans


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A novel isotopic labeling strategy is described for the structural analysis of proteins by NMR. Overexpression of a protein in a mammalian cell-line cultured in a medium containing amino acids labeled only in the backbone (N, Cα, Hα, C‘) atoms leads to the formation of exclusively backbone-labeled protein. We demonstrate that the absence of the one bond scalar coupling between the 13Cα and 13Cβ atoms that is observed in uniformly 13C enriched proteins offers a substantial sensitivity and resolution advantage in triple resonance NMR experiments that are commonly used to obtain backbone resonance assignments. This approach is illustrated in application to the β subunit of human chorionic gonadotropin isotopically enriched with 13C (97%), 15N (97%), and 2H (50%) exclusively in the backbone atoms of Phe, Val, and Leu residues.

Publication metadata

Author(s): Coughlin PE, Anderson FE, Oliver EJ, Brown JM, Homans SW, Pollak S, Lustbader JW

Publication type: Article

Publication status: Published

Journal: Journal of the American Chemical Society

Year: 1999

Volume: 121

Issue: 50

Pages: 11871-11874

Print publication date: 04/12/1999

ISSN (print): 0002-7863

ISSN (electronic): 1520-5126

Publisher: American Chemical Society


DOI: 10.1021/ja993083w


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