Investigation on the reaction conditions of Staphylococcus aureus sortase A for creating surface-modified liposomes as a drug-delivery system tool.

Atsushi Tabata, Natsuki Anyoji, Yukimasa Ohkubo, Toshifumi Tomoyasu, Hideaki Nagamune

Index: Anticancer Res. 34(8) , 4521-7, (2014)

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Abstract

This study aimed to determine the preferred conditions for the transpeptidase reaction of sortase A from Staphylococcus aureus, for the purpose of creating functional liposomes useful for a drug-delivery system (DDS).His-tagged recombinant sortase A with 59 amino acids deleted from the N-terminus (His-ΔN59SrtA) was prepared using an Escherichia coli expression system. The pH dependency and sorting signal sequence dependency of the transpeptidase reaction of His-ΔN59SrtA were analyzed by monitoring the transfer of model donor-substrates (i.e. His-tagged mutant green fluorescent proteins with a C-terminal LPxTG sorting signal) to model acceptor-beads with a GGGGGC peptide. In addition, using preferred conditions, the sortase A reaction was used to modify liposome surface.The transpeptidase reaction of His-ΔN59SrtA was enhanced under weakly acidic conditions. Transfer efficiency, based on sorting signal recognition by His-ΔN59SrtA, was similar to or higher than that obtained using several substrates with amino acids other than Glu in the sorting signal position "x". Furthermore, liposomes containing GGGGGC peptide-linked dipalmitoylphosphatidylethanolamine were successfully modified using the preferred conditions for His-ΔN59SrtA determined in this study.Preferred conditions for the transpeptidase reaction of His-ΔN59SrtA, especially in a weakly acidic environment to enhance reaction, was established and successfully used to create functional liposomes applicable to DDS.Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.