Collect. Czech. Chem. Commun. 2011, 76, 1089-1101
Published online 2011-08-22 09:30:38

Optimization of the posttranslational click modification of proteins

Milan Vrabel*, Emine Kaya, Stefan Prill, Veronika Ehmke and Thomas Carell*

Ludwig-Maximilian University Münich, Butenandtstrasse 5-13, D 81377 Münich, Germany


In order to develop efficient methods that would enable the synthesis of posttranslationaly modified proteins in a site-specific manner we have adopted the orthogonal pyrrolysyl-tRNA synthetase/tRNA pair to genetically encode various pyrrolysine analogs, which we were able to insert into the yellow fluorescent protein (YFP). These experiments showed that the alkene and alkyne containing amino acids 5 and 6 are superior substrates for the pyrrolysyl-tRNA synthetase and that they can be successfully incorporated into proteins. Using the Cu(I)-catalyzed Huisgen–Meldal–Sharpless click reaction, the alkyne containing YFP was finally glycosylated with various sugars. We confirmed the presence of the modified amino acids as well as the corresponding sugar modifications by HPLC-MS/MS mass spectrometry.

Keywords: Alkynes; Amino acids; Protein modification; Click chemistry; Triazoles; Azido compounds; Pyrrolysyl-tRNA synthetase; Pyrrolysine.

References: 32 live references.