8B6R

X-ray structure of the haloalkane dehalogenase HaloTag7 labeled with a chloroalkane Cyanine3 fluorophore substrate

Summary for 8B6R

• Entry DOI: 10.2210/pdb8b6r/pdb
• Related: 8B6S 8B6T
• Descriptor: Haloalkane dehalogenase, ~{N}-[2-[2-(6-chloranylhexoxy)ethoxy]ethyl]-6-[3,3-dimethyl-2-[(~{E})-3-(1,3,3-trimethylindol-2-ylidene)prop-1-enyl]indol-1-ium-1-yl]hexanamide, CHLORIDE ION, ... (6 entities in total)
• Functional Keywords: haloalkane dehalogenase, halotag, halotag7, self-labeling protein, fluorophore, cyanine 3, cy3, hydrolase
• Biological source: Rhodococcus sp.
• Total number of polymer chains: 1
• Total formula weight: 34317.47

Authors

Tarnawski, M.,Hellweg, L.,Hiblot, J. (deposition date: 2022-09-27, release date: 2023-07-26, Last modification date: 2024-11-20)

Primary citation

Hellweg, L.,Edenhofer, A.,Barck, L.,Huppertz, M.C.,Frei, M.S.,Tarnawski, M.,Bergner, A.,Koch, B.,Johnsson, K.,Hiblot, J.
A general method for the development of multicolor biosensors with large dynamic ranges.
Nat.Chem.Biol., 19:1147-1157, 2023

PubMed Abstract: Fluorescent biosensors enable the study of cell physiology with spatiotemporal resolution; yet, most biosensors suffer from relatively low dynamic ranges. Here, we introduce a family of designed Förster resonance energy transfer (FRET) pairs with near-quantitative FRET efficiencies based on the reversible interaction of fluorescent proteins with a fluorescently labeled HaloTag. These FRET pairs enabled the straightforward design of biosensors for calcium, ATP and NAD with unprecedented dynamic ranges. The color of each of these biosensors can be readily tuned by changing either the fluorescent protein or the synthetic fluorophore, which enables simultaneous monitoring of free NAD in different subcellular compartments following genotoxic stress. Minimal modifications of these biosensors furthermore allow their readout to be switched to fluorescence intensity, fluorescence lifetime or bioluminescence. These FRET pairs thus establish a new concept for the development of highly sensitive and tunable biosensors.

PubMed: 37291200
DOI: 10.1038/s41589-023-01350-1

Experimental method

X-RAY DIFFRACTION (1.5 Å)