The PinPoint Xa Protein Purification System is designed for the production and purification of fusion proteins that are biotinylated in vivo. The DNA coding for the protein of interest is cloned into a PinPoint Vector downstream of a sequence encoding a peptide that becomes biotinylated in vivo. Biotinylated fusion proteins are produced in E. coli and are affinity-purified using the SoftLink Soft Release Avidin Resin. This proprietary resin allows elution of the fusion protein under nondenaturing conditions. The PinPoint Vectors feature the encoded endoproteinase Factor Xa (pronounced ten a) proteolytic site that provides a way to separate the purification tag from the native protein, and the vectors carry a convenient multiple cloning region for ease in construction of fusion proteins. The system contains vectors in all possible sense reading frames, an avidin-conjugated resin, Streptavidin-Alkaline Phosphatase, a purification column and biotin. The PinPoint Xa Control Vector contains the chloramphenicol acetyltransferase (CAT) gene and is provided as a means of monitoring protein expression, purification and processing conditions. The system generally yields 1-5mg of protein per liter of culture.|
In vivo Biotinylation Tag: Allows purification of fusion proteins; many proteins produced have been soluble. Easy to Use: Purification of biotinylated proteins with the SoftLink™ Resin can be performed by column or batch purification. Easy Detection: Streptavidin Alkaline Phosphatase can be used to detect the biotinylated fusion protein in a pseudo-Western format to monitor purification. Flexible: PinPoint™Vectors are supplied for all reading frames. Gentle Release Conditions: SoftLink™ Resin allows release of the fusion protein under nondenaturing conditions. tac Promoter: Allows tightly regulated expression.