A genetically encodable microtag for chemo-enzymatic derivatization and purification of recombinant proteins

• Published in: Protein expression and purification 2005, Vol.39 (1), p.71-81
• Main Authors: Dursina, Beatrice-Elena, Reents, Reinhard, Niculae, Anca, Veligodsky, Alexei, Breitling, Reinhard, Pyatkov, Konstantin, Waldmann, Herbert, Goody, Roger S., Alexandrov, Kirill
• Format: Article
• Language: English
• Subjects: Electrophoresis, Polyacrylamide Gel; Monomeric GTP-Binding Proteins - isolation & purification; Monomeric GTP-Binding Proteins - metabolism; Phase partitioning; Protein prenylation; Protein Prenylation - physiology; Protein purification tags; Recombinant Proteins - isolation & purification; Recombinant Proteins - metabolism; Terpenes - chemical synthesis
• ISSN: 1046-5928; 1096-0279
• DOI: 10.1016/j.pep.2004.09.015

Efficient separation of recombinant polypeptides from proteins of the expression host and their subsequent derivatisation with functional chemical groups is essential for the success of many biological applications. Numerous tag systems have been developed to facilitate the purification procedure but only limited progress has been made in development of generic methods for targeted modification of proteins with functional groups. In this work, we present a novel 6 amino acid long C-terminal protein tag that can be selectively modified with functionalized derivatives of farnesyl isoprenoids by protein farnesyltransferase. The reaction could be performed in complex protein mixtures without detectable unspecific labeling. We demonstrate that this modification can be used to purify the target protein by over 800-fold in a single purification step using phase partitioning. Moreover, we show that the fluorescent group could be used to monitor the interaction of the derivatized proteins with other polypeptides.