One-Step Enzymatic Modification of the Cell Surface Redirects Cellular Cytotoxicity and Parasite Tropism

Surface display of engineered proteins
has many useful applications.
The expression of a synthetic chimeric antigen receptor composed of
an extracellular tumor-specific antibody fragment linked to a cytosolic
activating motif in engineered T cells is now considered a viable
approach for the treatment of leukemias. The risk of de novo tumor development, inherent in the transfer of genetically engineered
cells, calls for alternative approaches for the functionalization
of the lymphocyte plasma membrane. We demonstrate the conjugation
of LPXTG-tagged probes and LPXTG-bearing proteins to endogenous acceptors
at the plasma membrane in a single step using sortase A. We successfully
conjugated biotin probes not only to mouse hematopoietic cells but
also to yeast cells, 293T cells, and Toxoplasma gondii. Installation of single domain antibodies on activated CD8 T cell
redirects cell-specific cytotoxicity to cells that bear the relevant
antigen. Likewise, conjugation of Toxoplasma gondii with single domain antibodies targets the pathogen to cells that
express the antigen recognized by these single domain antibodies.
This simple and robust enzymatic approach enables engineering of the
plasma membrane for research or therapy under physiological reaction
conditions that ensure the viability of the modified cells.