This C-terminal Sortagging peptide acts as a (oligo)glycine nucleophile in the final steps of a sortagging protein labelling reaction. This reaction results in the [Lys(5-TAMRA)]- fluorescent moiety being attached to the C-terminus of the target protein or peptide.

A substrate peptide containing the LPXTG motif is recognised and cleaved by the enzyme Sortase A (SrtA) from Staphylococcus aureus. The catalytic cysteine residue in the active site of SrtA, serves as a nucleophile to cleave the peptide bond between threonine and glycine of the substrate peptide. Cleavage results in the formation of a thioacyl intermediate between the substrate peptide and SrtA. This intermediate is then resolved by the N-terminus of this (oligo)glycine nucleophile peptide, resulting in the creation of a new peptide bond that links the substrate peptide to this peptide and its fluorescent dye.  This method of protein labelling is known as sortagging.

5-Carboxytetramethylrhodamine (5-TAMRA) is a widely used fluorescent dye which excites at 546 nm and emits at 579 nm.