Routing Nanomolar Protein Cargoes to Lipid Raft-Mediated/Caveolar Endocytosis through a Ganglioside GM1-Specific Recognition Tag

Norbert Imre; Anasztázia Hetényi; Enikő Szabó; Brigitta Bodnár; Abel Szkalisity; Ilona Gróf; Alexandra Bocsik; Mária A Deli; Peter Horvath; Ágnes Czibula; Éva Monostori; Tamás A Martinek

doi:10.1002/advs.201902621

Routing Nanomolar Protein Cargoes to Lipid Raft-Mediated/Caveolar Endocytosis through a Ganglioside GM1-Specific Recognition Tag

Adv Sci (Weinh). 2020 Jan 9;7(4):1902621. doi: 10.1002/advs.201902621. eCollection 2020 Feb.

Authors

Norbert Imre¹, Anasztázia Hetényi¹, Enikő Szabó², Brigitta Bodnár^{1

3}, Abel Szkalisity⁴, Ilona Gróf^{5

6}, Alexandra Bocsik⁵, Mária A Deli^{5

7}, Peter Horvath⁸, Ágnes Czibula², Éva Monostori², Tamás A Martinek^{1

3}

Affiliations

¹ Department of Medical Chemistry University of Szeged Dóm tér 8 Szeged HU-6720 Hungary.
² Institute of Genetics Biological Research Center (BRC) Temesvári krt. 62 Szeged HU-6726 Hungary.
³ MTA-SZTE Biomimetic Systems Research Group University of Szeged Dóm tér 8 Szeged HU-6720 Hungary.
⁴ Synthetic and Systems Biology Unit Biological Research Center (BRC) Temesvári krt. 62 Szeged HU-6726 Hungary.
⁵ Institute of Biophysics Biological Research Center (BRC) Temesvári krt. 62 Szeged HU-6726 Hungary.
⁶ Doctoral School of Biology University of Szeged Dugonics tér 13 Szeged HU-6720 Hungary.
⁷ Department of Cell Biology and Molecular Medicine University of Szeged Somogyi u. 4 Szeged HU-6720 Hungary.
⁸ Institute for Molecular Medicine Finland University of Helsinki Tukholmankatu 8 Helsinki FI-00014 Finland.

Abstract

There is a pressing need to develop ways to deliver therapeutic macromolecules to their intracellular targets. Certain viral and bacterial proteins are readily internalized in functional form through lipid raft-mediated/caveolar endocytosis, but mimicking this process with protein cargoes at therapeutically relevant concentrations is a great challenge. Targeting ganglioside GM1 in the caveolar pits triggers endocytosis. A pentapeptide sequence WYKYW is presented, which specifically captures the glycan moiety of GM1 (K _D = 24 nm). The WYKYW-tag facilitates the GM1-dependent endocytosis of proteins in which the cargo-loaded caveosomes do not fuse with lysosomes. A structurally intact immunoglobulin G complex (580 kDa) is successfully delivered into live HeLa cells at extracellular concentrations ranging from 20 to 160 nm, and escape of the cargo proteins to the cytosol is observed. The short peptidic WYKYW-tag is an advantageous endocytosis routing sequence for lipid raft-mediated/caveolar cell delivery of therapeutic macromolecules, especially for cancer cells that overexpress GM1.

Keywords: cell delivery; endocytosis; glycan; immunoglobulin; peptides.