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Methyl side-chain dynamics in proteins using selective enrichment with a single isotopomer

Lookup NU author(s): Professor Steve Homans


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13C relaxation studies on side-chain methyl groups in proteins typically involve measurements on 13CHD2 isotopomers, where the 13C relaxation mechanism is particularly straightforward in the presence of a single proton. While such isotopomers can be obtained in proteins overexpressed in bacteria by use of 13C enriched and fractionally deuterated media, invariably all possible 2H isotopomers are obtained. This results in a loss of both resolution and sensitivity, which becomes particularly severe for larger proteins. We describe an approach that overcomes this problem by chemical synthesis of amino acids containing a pure 13CHD2 isotopomer. We illustrate the benefits of this approach in 13C side-chain relaxation measurements on the mouse major urinary protein selectively enriched with [γ1,γ2-13C2,α,β,γ1,γ1,γ2,γ2-2H6] valine. Relaxation measurements in the absence and presence of pyrazine-derived ligands suggest that valine side-chain dynamics do not contribute significantly to binding entropy.

Publication metadata

Author(s): Chaykovski MM, Bae LC, Cheng MC, Murray JH, Tortolani KE, Zhang R, Seshadri K, Findlay JBC, Hsieh SY, Homans SW, Kalverda AP, Brown JM

Publication type: Article

Publication status: Published

Journal: Journal of the American Chemical Society

Year: 2003

Volume: 125

Issue: 51

Pages: 15767-15771

ISSN (print): 0002-7863

ISSN (electronic): 1943-2984

Publisher: American Chemical Society


DOI: 10.1021/ja0368608


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