Analytic Selector: A rapid assay for detergent compatibility
Selector is a rapid, high throughput assay that explores detergent space to identify those best suited to stabilize the membrane protein of interest in advance of downstream protein crystallization trials. The assay uses differential filtration in 94 different detergents to generate information regarding both the stability and the size of the protein-detergent complex. The steps of this assay are simple: First, the protein is bound to an affinity resin, washed and eluted in a new detergent. The eluate is then passed through two filter plates with differing molecular weight cut-offs (MWCO). . The filtration profiles provide stability and relative sizing information that allows the selection of detergents most likely to stabilize a protein in a manner that would be commensurate with crystallization and structure determination.Why should I explore such a variety of detergents?
Integral membrane proteins present a unique challenge to protein chemists and structural biologists. In their natural environment, these proteins exist within a mosaic lipid bilayer that is a dynamic and complex environment. Prior to structural study, however, these must be extracted from the cell membrane and solubilized in such a way as to satisfy the hydrophobic nature of the transmembrane regions while ensuring that hydrophilic domains are in contact with an aqueous phase. While a significant number of detergents may be capable of keeping the protein in a soluble form, the stability, homogeneity and activity of any given membrane protein can be tremendously affected by the properties of the solubilizing component.
Deposits to the Protein Data Bank for membrane proteins comprise broad range of detergent and detergent-lipid complexes as part of the protein preparations leading the diffraction-quality crystals. Thus, while certain detergents (DM, DDM or OG, for example) are known for their success in stabilizing integral membrane proteins for structural studies and are reported at high frequency, significant effort would be required to explore detergent space, even to cover detergents previously successful for protein crystallization.
WHAT DOES THIS KIT CONTAIN?
1 x detergent screening plate containing 150 µl of 94 detergents at 2x working concentration, one blank and one position for the detergent currently stabilizing the protein sample to be screened
1 x 0.22 µm filter plate with receptacle plate for detergent exchange
1 x receptacle plate into which the proteins and newly exchanged detergents will be eluted
1 x 300 MWCO filter plate with matched receptacle plate
1 x 100 MWCO filter plate with matched receptacle plate.
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