NMR paper Reduced Dimensionality tailored HN(C)N Experiments for Facile Backbone Resonance Assignment of Proteins through Unambiguous Identification of Sequential HSQC Peaks

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[NMR paper] Reduced Dimensionality tailored HN(C)N Experiments for Facile Backbone Resonance Assignment of Proteins through Unambiguous Identification of Sequential HSQC Peaks

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10-09-2013, 05:31 AM

nmrlearner

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Reduced Dimensionality tailored HN(C)N Experiments for Facile Backbone Resonance Assignment of Proteins through Unambiguous Identification of Sequential HSQC Peaks

Reduced Dimensionality tailored HN(C)N Experiments for Facile Backbone Resonance Assignment of Proteins through Unambiguous Identification of Sequential HSQC Peaks

Publication date: Available online 8 October 2013
Source:Journal of Magnetic Resonance

Author(s): Dinesh Kumar

Two novel reduced dimensionality (RD) tailored HN(C)N [Panchal et. al., J. Biomol. NMR. (2002), 20 (2), 135-147] experiments are proposed to facilitate the backbone resonance assignment of proteins both in terms of its accuracy and speed. These experiments -referred here as (4,3)D-hNCOcaNH and (4,3)D-hNcoCANH- exploit the linear combination of backbone 15N and 13C?/13C? chemical shifts simultaneously to achieve higher peak dispersion and randomness along their respective F 1 dimensions. Simply, this has been achieved by modulating the backbone 15N(i) chemical shifts with that of 13C?(i-1)/13C? (i-1) spins following the established reduced dimensionality NMR approach [Szyperski et. al. (2002), Proc. Natl. Acad. Sci. U.S.A. 99(12), 8009-8014]. Though the modification is simple it has resulted an ingenious improvement of HN(C)N both in terms of peak dispersion and easiness of establishing the sequential connectivities. The increased dispersion along F 1 dimension solves two purposes here: (i) resolves the ambiguities arising because of degenerate 15N chemical shifts and (ii) reduces the signal overlap in F 2(15N)- F 3(1H) planes (an important requisite in HN(C)N based assignment protocol for facile and unambiguous identification of sequentially connected HSQC peaks). The performance of both these experiments and the assignment protocol has been demonstrated using bovine apo Calbindin-d9k (75 aa) and urea denatured UNC60B (a 152 amino acid ADF/cofilin family protein of Caenorhabditis elegans), as representatives of folded and unfolded protein systems, respectively.
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