(3,2)D GFT-NMR experiments for fast data collection from proteins

Youlin Xia; Guang Zhu; Sudha Veeraraghavan; Xiaolian Gao

doi:10.1023/B:JNMR.0000034352.75619.3f

(3,2)D GFT-NMR experiments for fast data collection from proteins

Youlin Xia, Guang Zhu, Sudha Veeraraghavan, Xiaolian Gao

Research output: Contribution to journal › Article › peer-review

21 Scopus citations

Abstract

High throughput structure determination of proteins will contribute to the success of proteomics investigations. The G-Matrix Fourier Transformation NMR (GFT-NMR) method significantly shortens experimental time by reducing the number of the dimensions of data acquisition for isotopically labeled proteins (Kim, S. and Szyperski, T.(2003) J. Am. Chem. Soc. 125, 1385). We demonstrate herein a suite of ten 3D\to2D or (3,2)D GFT-NMR experiments using ¹³C/¹⁵N-labeled ubiquitin. These experiments were completed within 18 hours, representing a 4- to 18-fold reduction in data acquisition time compared to the corresponding conventional 3D experiments. A subset of the GFT-NMR experiments, (3,2)D HNCO, HNCACB, HN(CO)CACB, and 2D ¹H-¹⁵N HSQC, which are necessary for backbone assignments, were carried out within 6 hours. To facilitate the analysis of the GFT-NMR spectra, we developed automated procedures for viewing and analyzing the GFT-NMR spectra. Our overall strategy allows (3,2)D GFT-NMR experiments to be readily performed and analyzed. Nevertheless, the increase in spectral overlap and the reduction in signal sensitivity in these fast NMR experiments presently limit their application to relatively small proteins.

Original language	English (US)
Pages (from-to)	467-476
Number of pages	10
Journal	Journal of biomolecular NMR
Volume	29
Issue number	4
DOIs	https://doi.org/10.1023/B:JNMR.0000034352.75619.3f
State	Published - Aug 2004
Externally published	Yes

Bibliographical note

Funding Information:
The 800 MHz NMR spectrometer at the University of Houston is funded by the W.M. Keck Foundation and the University of Houston. This research was supported by grants from NIH (GM49957) and the Robert A. Welch Foundation (E-1027). Dr Guang Zhu thanks the Hong Kong Research Grant Council for partial financial support. The authors are grateful to Dr Thomas D. Goddard (University of California, San Francisco) for the new improvements in SPARKY, and Dr David Cohen (University of Houston) for helpful discussions.

Keywords

GFT-NMR
Multidimensional NMR experiments
Protein structure determination
RD-NMR

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title = "(3,2)D GFT-NMR experiments for fast data collection from proteins",

abstract = "High throughput structure determination of proteins will contribute to the success of proteomics investigations. The G-Matrix Fourier Transformation NMR (GFT-NMR) method significantly shortens experimental time by reducing the number of the dimensions of data acquisition for isotopically labeled proteins (Kim, S. and Szyperski, T.(2003) J. Am. Chem. Soc. 125, 1385). We demonstrate herein a suite of ten 3D\to2D or (3,2)D GFT-NMR experiments using 13C/15N-labeled ubiquitin. These experiments were completed within 18 hours, representing a 4- to 18-fold reduction in data acquisition time compared to the corresponding conventional 3D experiments. A subset of the GFT-NMR experiments, (3,2)D HNCO, HNCACB, HN(CO)CACB, and 2D 1H-15N HSQC, which are necessary for backbone assignments, were carried out within 6 hours. To facilitate the analysis of the GFT-NMR spectra, we developed automated procedures for viewing and analyzing the GFT-NMR spectra. Our overall strategy allows (3,2)D GFT-NMR experiments to be readily performed and analyzed. Nevertheless, the increase in spectral overlap and the reduction in signal sensitivity in these fast NMR experiments presently limit their application to relatively small proteins.",

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author = "Youlin Xia and Guang Zhu and Sudha Veeraraghavan and Xiaolian Gao",

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(3,2)D GFT-NMR experiments for fast data collection from proteins

Abstract

Bibliographical note

Keywords

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