Lepre, C.A. & Moore, J.M. Microdrop screening: a rapid method to optimize solvent conditions for NMR spectroscopy of proteins. J. Biomol. NMR 12, 493-499

Vertex Pharmaceuticals, Inc., Cambridge, MA 02139-4242, USA.
Journal of Biomolecular NMR (Impact Factor: 3.14). 12/1998; 12(4):493-9. DOI: 10.1023/A:1008353000679
Source: PubMed


Determining appropriate solvent conditions is a crucial first step for carrying out NMR spectroscopy of proteins, but rapid and efficient methods for doing so are currently lacking. Microdrop screening examines a large number of different solvent conditions using very small amounts of protein and minimal labor. Starting from one initial buffer condition, small aliquots of protein solution are combined with an array of solutions in which concentration, pH, buffer type, and added stabilizers are systematically varied. The protein concentration of each microliter-sized test drop ('microdrop') is gradually changed using vapor diffusion, and the solubility of the protein is determined by visual examination. A variety of analytical techniques may be applied to the contents of the microdrops to monitor enzymatic activity, aggregation, ligand binding, and protein folding.

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    • "Microdrop screening was used to detect KIR2DS1 aggregation as described by Lepre and Moore [12]. We tested 23 different buffer conditions with a 24 well tissue culture plate and siliconized glass cover slips. 1 mL of each buffer was pipetted into each reservoir, and then, 2 µL aliquots of protein solution in starting buffer were pipetted onto the glass coverslips. 1 µL of each of reservoir buffer was added to each drop, and the solutions were mixed by gently drawing and expelling the solution in the pipette tip. "
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    PLoS ONE 08/2011; 6(8):e23052. DOI:10.1371/journal.pone.0023052 · 3.23 Impact Factor
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    • "Numerous efforts have been devoted to overcoming the solubility and sample stability issues. For example, extensive buffer screening (Bagby et al. 1997; Lepre and Moore 1998), addition of charged amino acids (Golovanov et al. 2004), or introduction of point mutants (Huang et al. 1996; Ito and Wagner 2004; Sun et al. 1999) have been successfully utilized to increase the solubility of the target proteins. However , these methods are often protein specific, largely based on trial and error, and may not be easily applicable to other systems. "
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