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Conformational distributions of unfolded polypeptides from novel NMR techniques
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10.1063/1.2838167
/content/aip/journal/jcp/128/5/10.1063/1.2838167
http://aip.metastore.ingenta.com/content/aip/journal/jcp/128/5/10.1063/1.2838167
View: Figures

Figures

Image of FIG. 1.
FIG. 1.

Backbone conformations in proteins. Top: Due to the planarity of the peptide bond, only two angles and determine the backbone conformation of amino acids in proteins. Atoms are shown in CPK colors with hydrogen in white, nitrogen in blue, carbon in gray, and oxygen in red. Bottom: Ramachandran plot of and , where each point represents one amino acid conformation. 15 000 conformations are shown. The data points were randomly drawn from the protein coil library (Ref. 20) and correspond to nonglycine residues in the coil regions of protein crystal structures. The highest populated amino acid conformational basins are labeled as -extended , polyproline II (PPII), right-handed -helix , and left-handed -helix .

Image of FIG. 2.
FIG. 2.

Residual order of a bond vector (NH) in an elongated unfolded protein oriented by steric alignment in an elongated cavity that is parallel to the magnetic field. The depicted situation corresponds to radially squeezed gels. Extended conformations (right top) will contribute negative RDCs to the ensemble average, as the NH vectors are predominantly orthogonal to the magnetic field and the integral over is negative [Eq. (3)]. In contrast, NH vectors are parallel to the helix axis in helical conformations. For longitudinal alignment of the helices, the NH vector mostly samples directions parallel to the magnetic field, causing an inversion of the sign of RDCs from the dependence on . The predominance of extended conformations in highly unfolded proteins leads to mostly negative RDCs in radially squeezed gels.

Image of FIG. 3.
FIG. 3.

RDCs for a random flight chain model of the unfolded polypetide chain. Profiles are obtained from an analytical description for chains of length 11, 21, 51, and 101. [Data redrawn with permission from O. I. Obolensky, K. Schlepckow, H. Schwalbe, and A. V. Solov’yov, J. Biomol. NMR39, 1 (2007). Copyright 2007, Journal of Biomolecular NMR.]

Image of FIG. 4.
FIG. 4.

Detection of long-range order in -synuclein by RDCs. (A) Experimental data (solid, red) obtained by alignment in Pf1 bacteriophages under high salt concentrations. Under these conditions, alignment is mainly steric. Simulated data (dashed, blue) were obtained by the FM approach without long-range contacts. (B) Experimental data (solid, red) as in (A). Simulated data (dashed, blue) were obtained by FM with the additional constraint of long-range contacts between residue regions 1-20 and 121-140. [Data are taken with permission from P. Bernado, C. W. Bertoncini, C. Griesinger, M. Zweckstetter, and M. Blackledge, J. Am. Chem. Soc.127, 17968 (2005). Copyright 2005, Journal of the American Chemical Society.]

Image of FIG. 5.
FIG. 5.

and RDCs in the T4 fibritin foldon hairpin between 40 and at 2. Averages of the and RDCs at are shown as dotted lines. RDCs serve to directly monitor the order-disorder transition in the structured hairpin region (residues 12-16 and 19-23, indicated on top) during thermal unfolding. [Figure adapted with permission from S. Meier, S. Guthe, T. Kiefhaber, and S. Grzesiek, J. Mol. Biol.344, 1051 (2004). Copyright 2004, Journal of Molecular Biology.]

Image of FIG. 6.
FIG. 6.

Comparison of experimental (solid line, red) and predicted (dashed line, blue) RDCs in urea-denatured staphylococcal nuclease mutant . RDCs are predicted by the coil model using sidechain steric exclusion. [Figure adapted with permission from P. Bernado, L. Blanchard, P. Timmins, D. Marion, R. W. Ruigrok, and M. Blackledge, Proc. Natl. Acad. Sci. U.S.A.102, 17002 (2005). Copyright 2005, Proceedings of the National Academy Sciences of the United States of America.]

Image of FIG. 7.
FIG. 7.

Left: Strand of -sheet conformation showing an extended set of RDCs measured to refine the conformational model of urea-denatured ubiquitin. Middle, right: Agreement of the observed RDCs (solid, red) with predictions (dashed, blue) from the coil model (middle) and from a modified coil model assuming increased sampling of extended backbone conformational basins under denaturing conditions (right). Values of indicate additionally necessary scaling factors. [Figure adapted with permission from S. Meier, S. Grzesiek, and M. Blackledge, J. Am. Chem. Soc.129, 9799 (2007). Copyright 2007, Journal of the American Chemical Society.]

Image of FIG. 8.
FIG. 8.

Top: Conformational distributions of four sequential amino acids in denatured ubiquitin derived by increased sampling of extended backbone conformational basins in order to optimally reproduce measured multiple RDCs (Fig. 7). Bottom: Comparison to the database sampling of the unbiased coil model. Red indicates higher populations, yellow intermediate populations, and blue lower populations. [Figure reprinted with permission from S. Meier, S. Grzesiek, and M. Blackledge, J. Am. Chem. Soc.129, 9799 (2007). Copyright 2007, Journal of the American Chemical Society.]

Image of FIG. 9.
FIG. 9.

Detection of hydrogen bonds in ubiquitin under highly denaturing conditions ( urea). Top: strips from a three-dimensional long-range HNCO-TROSY experiment showing correlations across H bonds. Peaks are labeled with assignment information. Hydrogen bond correlations are marked in magenta. Bottom: Summary of long-range interactions in the first hairpin of urea-denatured ubiquitin. Detected interactions are shown as dashed cylinders for (magenta) and (gold). Ambiguous correlations due to spectral overlap are shown in white. [Figure reprinted with permission from S. Meier, M. Strohmeier, M. Blackledge, and S. Grzesiek, J. Am. Chem. Soc.129, 754 (2007). Copyright 2007, Proceedings of the National Academy of Sciences of the United States of America].

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2008-02-04
2014-04-16
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752b84549af89a08dbdd7fdb8b9568b5 journal.articlezxybnytfddd
Scitation: Conformational distributions of unfolded polypeptides from novel NMR techniques
http://aip.metastore.ingenta.com/content/aip/journal/jcp/128/5/10.1063/1.2838167
10.1063/1.2838167
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