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J Biomol NMR. 2015 Jan;61(1):47-53. doi: 10.1007/s10858-014-9881-8. Epub 2014 Nov 27.

APSY-NMR for protein backbone assignment in high-throughput structural biology.

Author information

1
Department of Integrative Structural and Computational Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA, 92037, USA.

Abstract

A standard set of three APSY-NMR experiments has been used in daily practice to obtain polypeptide backbone NMR assignments in globular proteins with sizes up to about 150 residues, which had been identified as targets for structure determination by the Joint Center for Structural Genomics (JCSG) under the auspices of the Protein Structure Initiative (PSI). In a representative sample of 30 proteins, initial fully automated data analysis with the software UNIO-MATCH-2014 yielded complete or partial assignments for over 90 % of the residues. For most proteins the APSY data acquisition was completed in less than 30 h. The results of the automated procedure provided a basis for efficient interactive validation and extension to near-completion of the assignments by reference to the same 3D heteronuclear-resolved [(1)H,(1)H]-NOESY spectra that were subsequently used for the collection of conformational constraints. High-quality structures were obtained for all 30 proteins, using the J-UNIO protocol, which includes extensive automation of NMR structure determination.

PMID:
25428764
PMCID:
PMC4305044
DOI:
10.1007/s10858-014-9881-8
[Indexed for MEDLINE]
Free PMC Article

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