HaloTag® Protein Purification System

The HaloTag® Protein Purification System is designed to purify proteins fused to HaloTag®, a novel protein tag that enhances the expression and solubility of recombinant proteins. HaloTag® Technology enables the covalent, efficient and specific capture of a protein of interest onto HaloLink™ Resin, thus overcoming the equilibrium-based limitations associated with affinity tags (i.e., poor capture of proteins expressed at low levels and protein loss during washing of the purification resin.

The HaloTag® technology offers a quick and convenient way to test protein expression of HaloTag® fusio...

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The HaloTag® Protein Purification System is designed to purify proteins fused to HaloTag®, a novel protein tag that enhances the expression and solubility of recombinant proteins. HaloTag® Technology enables the covalent, efficient and specific capture of a protein of interest onto HaloLink™ Resin, thus overcoming the equilibrium-based limitations associated with affinity tags (i.e., poor capture of proteins expressed at low levels and protein loss during washing of the purification resin.

The HaloTag® technology offers a quick and convenient way to test protein expression of HaloTag® fusion proteins as well as monitor the efficiency of immobilization to HaloLink™ Resin by labeling with fluorescent HaloTag® TMR Ligand followed by SDS-PAGE analysis. Guidelines can be found in the HaloLink™ Resin Technical Manual #TM250, the HaloLink™ Protein Array Technical Manual #TM310 and the HaloCHIP™ System Technical Manual #TM075.

Outline of Procedure
The HaloTag® protein, a 34kDa mutated hydrolase, covalently attaches to HaloLink™ Resin via an immobilized chloroalkane ligand. TEV Protease cleaves the target protein from the HaloLink™ Resin. The TEV Protease, which has an N-terminal (HQ) tag, is removed from the protein of interest using HisLink™ Resin, and the purified protein of interest is recovered. The appropriate vector that encodes the HaloTag® protein and expresses protein optimally in E. coli is pFN18A HaloTag® T7 Flexi® Vector (G2751) or pFN18K HaloTag® T7 Flexi® Vector (G2681). These vectors can be purchased separately.

Features - Benefits

• Experience Superior Yield, Purity and Specific Activity of Soluble, Functional Proteins Compared to His-Tag, GST and MBP Affinity Tags: Specific and covalent HaloTag® fusion protein capture and immobilization on HaloLink™ Resin.
• Achieve Enhanced Target Protein Expression in Prokaryotic, Mammalian and Cell-Free Systems: Proteins are expressed as HaloTag® fusion proteins.
• Purify Poorly Expressed Fusion Proteins: Rapid, specific and covalent capture of HaloTag® protein onto HaloLink™ Resin is a nonequilibrium process.
• Efficiently Recover Tag-Free Target Protein using TEV Protease Cleavage: Optimized TEV protease recognition site within the interconnecting polypeptide separating the HaloTag® protein and the fusion partner. HaloTag® protein remains immobilized on the resin due to covalent capture.
• Save Time: One buffer compatible with downstream applications for all purification steps.
• Perform Easy In-Gel Detection and Quantification of Protein Expression Levels with Fluorescent HaloTag® Ligands: Highly stable HaloTag® protein-ligand interaction permits boiling with SDS sample buffer followed by resolving on SDS-PAGE.

Applications

• Purification of HaloTag® fusion proteins expressed in E. coli, especially for proteins that are otherwise difficult to express.
• Purification of HaloTag® fusion proteins expressed in mammalian and cell-free systems.

Notes

Cat.# G6280 contains 100ml of a 25% slurry of HaloLink™ Resin, TEV Protease and HisLink™ Resin. Cat.# G6270 contains 10ml of a 25% slurry of HaloLink™ Resin, TEV Protease and HisLink™ Resin.

Competitive pricing is available for bulk quantities of HaloLink™ Resin (e.g., more than 100 ml). Please inquire.

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