title: Development of a dehalogenase-based protein fusion tag capable of rapid, selective and covalent attachment to customizable ligands.
abstract: Our fundamental understanding of proteins and their biological significance has been enhanced by genetic fusion tags, as they provide a convenient method for introducing unique properties to proteins so that they can be examinedin isolation. Commonly used tags satisfy many of the requirements for applications relating to the detection and isolation of proteins from complex samples. However, their utility at low concentration becomes compromised if the binding affinity for a detection or capture reagent is not adequate to produce a stable interaction. Here, we describe HaloTag® (HT7), a genetic fusion tag based on a modified haloalkane dehalogenase designed and engineered to overcome the limitation of affinity tags by forming a high affinity, covalent attachment to a binding ligand. HT7 and its ligand have additional desirable features. The tag is relatively small, monomeric, and structurally compatible with fusion partners, while the ligand is specific, chemically simple, and amenable to modular synthetic design. Taken together, the design features and molecular evolution of HT7 have resulted in a superior alternative to common tags for the overexpression, detection, and isolation of target proteins.
authors.affiliation: Promega Corporation, Madison, WI, USA.
authors.forename: Lance P
authors.initials: LP
authors.lastname: Encell
authors.forename: Rachel
authors.initials: R
authors.lastname: Friedman Ohana
authors.forename: Kris
authors.initials: K
authors.lastname: Zimmerman
authors.forename: Paul
authors.initials: P
authors.lastname: Otto
authors.forename: Gediminas
authors.initials: G
authors.lastname: Vidugiris
authors.forename: Monika G
authors.initials: MG
authors.lastname: Wood
authors.forename: Georgyi V
authors.initials: GV
authors.lastname: Los
authors.forename: Mark G
authors.initials: MG
authors.lastname: McDougall
authors.forename: Chad
authors.initials: C
authors.lastname: Zimprich
authors.forename: Natasha
authors.initials: N
authors.lastname: Karassina
authors.forename: Randall D
authors.initials: RD
authors.lastname: Learish
authors.forename: Robin
authors.initials: R
authors.lastname: Hurst
authors.forename: James
authors.initials: J
authors.lastname: Hartnett
authors.forename: Sarah
authors.initials: S
authors.lastname: Wheeler
authors.forename: Pete
authors.initials: P
authors.lastname: Stecha
authors.forename: Jami
authors.initials: J
authors.lastname: English
authors.forename: Kate
authors.initials: K
authors.lastname: Zhao
authors.forename: Jacqui
authors.initials: J
authors.lastname: Mendez
authors.forename: Hélène A
authors.initials: HA
authors.lastname: Benink
authors.forename: Nancy
authors.initials: N
authors.lastname: Murphy
authors.forename: Danette L
authors.initials: DL
authors.lastname: Daniels
authors.forename: Michael R
authors.initials: MR
authors.lastname: Slater
authors.forename: Marjeta
authors.initials: M
authors.lastname: Urh
authors.forename: Aldis
authors.initials: A
authors.lastname: Darzins
authors.forename: Dieter H
authors.initials: DH
authors.lastname: Klaubert
authors.forename: Robert F
authors.initials: RF
authors.lastname: Bulleit
authors.forename: Keith V
authors.initials: KV
authors.lastname: Wood