Chemoenzymatic Labeling to Visualize Intercellular Contacts Using Lipidated SortaseA**

Abstract

Methods to label intercellular contact have attracted attention because of their potential in cell biological and medical applications for the analysis of intercellular communications. In this study, a simple and versatile method for chemoenzymatic labeling of intercellularly contacting cells is demonstrated using a cell‐surface anchoring reagent of a poly(ethylene glycol)(PEG)‐lipid conjugate. The surface of each cell in the cell pairs of interest were decorated with sortase A (SrtA) and triglycine peptide that were lipidated with PEG‐lipid. In the mixture of the two‐cell populations, the triglycine‐modified cells were enzymatically labeled with a fluorescent labeling reagent when in contact with SrtA‐modified cells on a substrate. The selective labeling of the contacting cells was confirmed by confocal microscopy. The method is a promising tool for selective visualization of intercellularly contacting cells in cell mixtures for cell‐cell communication analysis.

Promiscuous Enzymes for Residue‐Specific Peptide and Protein Late‐Stage Functionalization

“…A directed sortase evolution strategy has been used to generate variants with up to 22‐fold increase in catalytic efficiency compared to the wild‐type enzyme. SrtA was also utilized to probe intracellular contact using a PEG‐lipid conjugate to a triglycine peptide followed by fluorescence labelling [126] . It has been shown to have broad substrate tolerance allowing it to modify peptidoglycan as well as its precursor lipid II [127] .…”

“…SrtA was also utilized to probe intracellular contact using a PEG-lipid conjugate to a triglycine peptide followed by fluorescence labelling. [ 126 ] It has been shown to have broad substrate tolerance allowing it to modify peptidoglycan as well as its precursor lipid II. [ 127 ] SrtA has found applications in protein modifications and immobilization on surfaces and synthesis of protein polymer conjugates.…”