Paramagnetic lanthanide tags for site-specific ligation to an unnatural amino acid to generate pseudocontact shifts in proteins — ASN Events
  1. Schedule
  2. Session Ten - Poster Session C
  3. Paramagnetic lanthanide tags for site-specific ligation to an unnatural amino acid to generate pseudocontact shifts in proteins

Paramagnetic lanthanide tags for site-specific ligation to an unnatural amino acid to generate pseudocontact shifts in proteins (#348)

Choy Theng Loh 1 , Kiyoshi Ozawa 1 , Kellie L. Tuck 2 , Nicholas Barlow 3 , Thomas Huber 1 , Gottfried Otting 1 , Bim Graham 3
  1. Research School of Chemistry, The Australian National University, Acton, ACT, Australia
  2. School of Chemistry, Monash University, Clayton, VIC, Australia
  3. Medicinal Chemistry, Monash Institute of Pharmaceutical Sciences, Parkville, VIC, Australia

Paramagnetic labelling of proteins is becoming an increasingly popular tool for structural biology by NMR spectroscopy. Pseudocontact shifts (PCSs) induced by paramagnetic lanthanide ions provide particularly valuable long-range structural information. The challenge is to attach the lanthanide both site-specifically and in a rigid manner, as even limited mobility of the lanthanide ion relative to the protein can average the observed PCSs to zero. Our approach is to incorporate an unnatural amino acid into the protein using an orthogonal tRNA/aminoacyl-tRNA synthetase pair [1]. We successfully incorporated p-azido-L-phenyalanine into different proteins and established conditions for high-yield ligation of different lanthanide-binding tags by copper-catalyzed azide-alkyne cycloaddition. PCSs were reliably generated by this approach.