6Y7B

X-ray structure of the Haloalkane dehalogenase HaloTag7 labeled with a chloroalkane-carbopyronine fluorophore substrate

Summary for 6Y7B

• Entry DOI: 10.2210/pdb6y7b/pdb
• Related: 6Y7A
• Descriptor: Haloalkane dehalogenase, 4-[2-[2-(6-chloranylhexoxy)ethoxy]ethylcarbamoyl]-2-[3-(dimethylamino)-6-(dimethyl-$l^{4}-azanylidene)-10,10-dimethyl-anthracen-9-yl]benzoic acid, CHLORIDE ION (3 entities in total)
• Functional Keywords: haloalkane dehalogenase, halo, tag, halotag7, self-labeling protein, fluorophore, carbopyronine, hydrolase
• Biological source: Rhodococcus sp.
• Total number of polymer chains: 5
• Total formula weight: 169623.50

Authors

Tarnawski, M.,Johnsson, K.,Hiblot, J. (deposition date: 2020-02-28, release date: 2021-03-31, Last modification date: 2024-10-16)

Primary citation

Wilhelm, J.,Kuhn, S.,Tarnawski, M.,Gotthard, G.,Tunnermann, J.,Tanzer, T.,Karpenko, J.,Mertes, N.,Xue, L.,Uhrig, U.,Reinstein, J.,Hiblot, J.,Johnsson, K.
Kinetic and Structural Characterization of the Self-Labeling Protein Tags HaloTag7, SNAP-tag, and CLIP-tag.
Biochemistry, 60:2560-2575, 2021

PubMed Abstract: The self-labeling protein tags (SLPs) HaloTag7, SNAP-tag, and CLIP-tag allow the covalent labeling of fusion proteins with synthetic molecules for applications in bioimaging and biotechnology. To guide the selection of an SLP-substrate pair and provide guidelines for the design of substrates, we report a systematic and comparative study of the labeling kinetics and substrate specificities of HaloTag7, SNAP-tag, and CLIP-tag. HaloTag7 reaches almost diffusion-limited labeling rate constants with certain rhodamine substrates, which are more than 2 orders of magnitude higher than those of SNAP-tag for the corresponding substrates. SNAP-tag labeling rate constants, however, are less affected by the structure of the label than those of HaloTag7, which vary over 6 orders of magnitude for commonly employed substrates. Determining the crystal structures of HaloTag7 and SNAP-tag labeled with fluorescent substrates allowed us to rationalize their substrate preferences. We also demonstrate how these insights can be exploited to design substrates with improved labeling kinetics.

PubMed: 34339177
DOI: 10.1021/acs.biochem.1c00258

Experimental method

X-RAY DIFFRACTION (3.1 Å)