[NMR paper] NMR screening and crystal quality of bacterially expressed prokaryotic and eukaryotic

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NMR screening and crystal quality of bacterially expressed prokaryotic and eukaryotic proteins in a structural genomics pipeline.

Related Articles NMR screening and crystal quality of bacterially expressed prokaryotic and eukaryotic proteins in a structural genomics pipeline.

Proc Natl Acad Sci U S A. 2005 Feb 8;102(6):1901-5

Authors: Page R, Peti W, Wilson IA, Stevens RC, Wüthrich K

In the Joint Center for Structural Genomics, one-dimensional (1D) 1H NMR spectroscopy is routinely used to characterize the folded state of protein targets and, thus, serves to guide subsequent crystallization efforts and to identify proteins for NMR structure determination. Here, we describe 1D 1H NMR screening of a group of 79 mouse homologue proteins, which correlates the NMR data with the outcome of subsequent crystallization experiments and crystallographic structure determination. Based on the 1D 1H NMR spectra, the proteins are classified into four groups, "A" to "D." A-type proteins are candidates for structure determination by NMR or crystallography; "B"-type are earmarked for crystallography; "C" indicates folded globular proteins with broadened line shapes; and "D" are nonglobular, "unfolded" polypeptides. The results obtained from coarse- and fine-screen crystallization trials imply that only A- and B-type proteins should be used for extensive crystallization trials in the future, with C and D proteins subjected only to coarse-screen crystallization trials. Of the presently studied 79 soluble protein targets, 63% yielded A- or B-quality 1D 1H NMR spectra. Although similar yields of crystallization hits were obtained for all four groups, A to D, crystals from A- and B-type proteins diffracted on average to significantly higher resolution than crystals produced from C- or D-type proteins. Furthermore, the output of refined crystal structures from this test set of proteins was 4-fold higher for A- and B-type than for C- and D-type proteins.

PMID: 15677718 [PubMed - indexed for MEDLINE]



Source: PubMed