Products Cited in Publications Worldwide Genetic Encoding of 7-Aza-L-tryptophan: Isoelectronic Substitution of a Single CH-Group in a Protein for a Nitrogen Atom for Site-Selective Isotope Labeling

Genetic Encoding of 7-Aza-L-tryptophan: Isoelectronic Substitution of a Single CH-Group in a Protein for a Nitrogen Atom for Site-Selective Isotope Labeling

Abdelkader, Elwy H.; Qianzhu, Haocheng; Huber, Thomas; Otting, Gottfried

DOI: PMID:

Abstract

Genetic encoding of a noncanonical amino acid (ncAA) in an in vivo expression system requires an aminoacyl-tRNA synthetase that specifically recognizes the ncAA, while the ncAA must not be recognized by the canonical protein expression machinery. We succeeded in genetically encoding 7-aza-tryptophan (7AW), which is isoelectronic with tryptophan. The system is fully orthogonal to protein expression in E. coli, enabling high-yielding site-selective isotope-labeling in vivo. 7AW is readily synthesized from serine and 7-aza-indole using a tryptophan synthetase β-subunit (TrpB) mutant, affording easy access to isotope-labeled 7AW. Using labeled 7AW produced from 15N/13C-labeled serine, we produced 7AW mutants of the 25 kDa Zika virus NS2B-NS3 protease. 15N-HSQC spectra display single cross-peaks at chem. shifts near those observed for the wild-type protein labeled with 15N/13C-tryptophan, confirming the structural integrity of the protein and yielding straightforward NMR resonance assignments for site-specific probing.

Keywords

7-azatryptophan ; genetic encoding ; isoelectronicsubstitution ; NMR spectroscopy ; selective isotopelabeling

Purchased from AmBeed

7-Azaindole

A156097271-63-698%

7-Azaindole

12.0/10g

Detail