Site-Specific Protein Modification on Living Cells Catalyzed by Sortase

✍ Scribed by Tsutomu Tanaka; Teruyasu Yamamoto; Shinya Tsukiji; Teruyuki Nagamune

• Book ID: 102788702
• Publisher: John Wiley and Sons
• Year: 2008
• Tongue: English
• Weight: 738 KB
• Volume: 9
• Category: Article
• ISSN: 1439-4227
• DOI: 10.1002/cbic.200700614

✦ Synopsis

Abstract

The use of enzymes is a promising approach for site‐specific protein modification on living cells owing to their substrate specificity. Herein we describe a general strategy for the site‐specific modification of cell surface proteins with synthetic molecules by using Sortase, a transpeptidase from Staphylococcus aureus. The short peptide tag LPETGG is genetically introduced to the C terminus of the target protein, expressed on the cell surface. Subsequent addition of Sortase and an N‐terminal triglycine‐containing probe results in the site‐specific labeling of the tagged protein. We were successful in the C‐terminal‐specific labeling of osteoclast differentiation factor (ODF) with a biotin‐ or fluorophore‐containing short peptide on the living cell surface. The labeling reaction occurred efficiently in serum‐containing medium, as well as serum‐free medium or PBS. The labeled products were detected after incubation for 5 min. In addition, site‐specific protein–protein conjugation was successfully demonstrated on a living cell surface by the Sortase‐catalyzed reaction. This strategy provides a powerful tool for cell biology and cell surface engineering.