HaloTag labeling {technology|technologies} has introduced unrivaled {potential|possible|prospective} in protein chemistry, molecular and cellular biology. A wide {variety of|number of|selection of} ligands {have been|happen to be|have already been} {developed|created} to meet the {specific|particular|certain|distinct|precise} {needs|requirements|wants|demands|desires|requires} of diverse applications, but only a single protein tag, DhaAHT, is routinely {used|utilized|employed|utilised|applied|made use of} for their incorporation. Following a systematic kinetic and computational {analysis|evaluation} of {different|various|distinct|diverse|unique|distinctive} reporters, tetramethylrhodamine and {three|3} 4-stilbazolium-based fluorescent ligands, we showed that the mechanism of incorporating {different|various|distinct|diverse|unique|distinctive} ligands depends {both|each} {on the|around the} binding step {and the|and also the|as well as the|along with the|plus the} efficiency {of the|from the|in the|on the|with the|of your} chemical reaction. By studying the {different|various|distinct|diverse|unique|distinctive} haloalkane dehalogenases DhaA, LinB, and DmmA, we {found|discovered|identified|located} that the architecture {of the|from the|in the|on the|with the|of your} access tunnels is {critical|crucial|vital|essential|important} for the kinetics of {both|each} {steps|actions|measures|methods} {and the|and also the|as well as the|along with the|plus the} ligand specificity. We show that {highly|extremely|very|hugely} {efficient|effective} labelling with {specific|particular|certain|distinct|precise} ligands is achievable with {natural|all-natural|organic} dehalogenases. We propose a {simple|easy|straightforward|basic|uncomplicated|very simple} protocol for {selecting|choosing|picking|deciding on} the optimal protein tag {for a|to get a|for any} {specific|particular|certain|distinct|precise} ligand from a wide pool of {available|accessible|obtainable|offered|readily available|out there} enzymes with diverse access tunnel architectures. The application of this protocol eliminates a {need|require|want|need to have|will need|have to have} for {expensive|costly|pricey|high-priced|high priced|highly-priced} and laborious protein engineering. Formula of N-(3-Chloro-4-hydroxyphenyl)acetamide Mal-PEG3-NHS ester site PMID:23399686
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