HaloTag7 self-labeling protein tag
HaloTag7, a catalytically inactive haloalkane dehalogenase derivative that forms a rapid, essentially irreversible covalent bond with synthetic chloroalkane ligands. The same genetic construct can be fluorescently labeled, captured on resin or immobilized, depending on the functional group on the ligand, and it enhances soluble expression in E. coli.
Origin: Engineered derivative of Rhodococcus haloalkane dehalogenase DhaA
Characteristics
Encodes a 296-amino-acid protein of 888 nucleotides, beginning AEIGTGFP and ending LSTLEISG. HaloTag7 is a 34 kDa monomer with a negative surface charge (pI about 5). Labeling is 1:1 stoichiometric, and the bond survives SDS-PAGE. As an N-terminal fusion in E. coli, it kept 74% of 23 human proteins soluble, versus 52% for MBP, 39% for GST and 22% for His6. This record is the tag without a protease site; Halo-TEV adds one.
Applications: Covalent fluorescent labeling and in-gel quantitation of fusion proteins in lysate, one-step capture on HaloLink resin with TEV release of the partner, immobilization on slides, soluble expression of human proteins in E. coli, and live and fixed cell imaging with fluorescent chloroalkane ligands.
Limitations: At 34 kDa it is larger than GST (26 kDa) and His6, and a tag can interfere with the structure or function of a partner. The covalent bond is irreversible, so a captured fusion cannot be eluted from the resin and is released only if a TEV site separates tag and partner. A chloroalkane ligand must be supplied separately.
Sequence
References
- Ohana et al. (2009). HaloTag7: A genetically engineered tag that enhances bacterial expression of soluble proteins and improves protein purification. Protein Expr Purif - Ohana 2009 HaloTag7