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A simple apparatus for generating stretched polyacrylamide gels, yielding uniform alignment of proteins and detergent micelles*

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Abstract

Compressed and stretched polyacrylamide hydrogels previously have been shown to offer a robust method for aligning proteins. A simple, funnel-like apparatus is described for generating uniformly stretched hydrogels. For prolate-shaped proteins, gels stretched in the direction of the magnetic field yield two-fold larger alignment than gels compressed to the same aspect ratio in this direction. Empirically, protein alignment is found to be proportional to (c−2.3)2 [(do/dN)3-1], where do and dN are the diameters of the cylindrical gels before and after stretching, respectively, and c is the polyacrylamide weight fraction in percent. Low gel densities, in the 4–7% range, are found to have minimal effects on macromolecular rotational correlation times, τc, and no effect of the compression ratio on τc could be discerned over the range studied (do/dN le1.4). Application is demonstrated for a sample containing the first Ig-binding domain of protein G, and for a detergent-solubilized peptide.

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Authors and Affiliations

  1. Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland, 20892-0520, U.S.A

    James J. Chou, Sander Gaemers, Bernard Howder, John M. Louis & Ad Bax

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  1. James J. Chou
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  2. Sander Gaemers
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  3. Bernard Howder
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  4. John M. Louis
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  5. Ad Bax
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Chou, J.J., Gaemers, S., Howder, B. et al. A simple apparatus for generating stretched polyacrylamide gels, yielding uniform alignment of proteins and detergent micelles*. J Biomol NMR 21, 377–382 (2001). https://doi.org/10.1023/A:1013336502594

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  • DOI: https://doi.org/10.1023/A:1013336502594

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