Detection of Early-Stage Protein Crystallization Using DPI A. Boudjemline a , N.J. Freeman a , D.T. Clarke b , G.R. Jones b . a Farfield Sensors Ltd, Unit 51 Salford Business Park, Leslie Hough Way, Manchester, M6 6AJ, United Kingdom. b CCLRC Daresbury Laboratory, Keckwick Lane, Warrington, WA4 4AD, United Kingdom

Introduction The three-dimensional structure of a protein, which is used to deduce its biological function necessary for drug design, is presently mainly determined by X-ray crystallography. However, the growth of suitable crystals has proved to be a major bottleneck as proteins are extremely hard to crystallize. Each protein has its own specific set of crystallization conditions, and no generic rule of crystallization exists. As a result, hundreds, or even thousands, of crystallization trials must be performed on a target protein, of which less than 1% typically yields promising results. At Farfield, we have conducted protein crystallization experiments using the Dual Polarization Interferometry (DPI) technique. The results using standard proteins, such as lysozyme, betalactoglobulin, myoglobin and glucose isomerase, showed that the technique can be very useful and effective in detecting the earliest stages of protein crystallization. Technique The DPI technique, as deployed in Farfield's Ana light ? BIO200 instruments, consists of a dual slab waveguide sensor chip illuminated with an alternating polarised laser beam. The chip works as an integrated Young's interferometer generating interference fringes at its output as depicted in Figure 1a (bottom waveguide: reference, and top waveguide: sensing layer). Changes in the refractive index (RI) at the surface of the chip will manifest as a phase shift in the exiting fringes. This phase shift is directly related to the RI and thickness of any layer (e.g., protein) that adsorbs to the surface. By exciting the chip with two orthogonal polarizations, i.e., TE and TM, two separate measurements are made which can be unambiguously resolved into the thickness and RI of the layer. The generated fringe pattern is characterised by a few parameters, including its period and contrast as defined in Figure 1b. The contrast is a measure of the amount of light guided in both waveguides, and as such, is affected by any losses, e.g., scattering that occurs at the top active waveguide. It is this property that we used to monitor protein crystallization.

Experimental

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Results 1 .Lysozyme crystallization

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