Direct Peptide Interaction with Surface Glycosaminoglycans Contributes to the Cell Penetration of Maurocalcine

Small Efficient Cell-penetrating Peptides Derived from Scorpion Toxin Maurocalcine

“…Because amiloride is an exquisite blocker of macropinosomes (22)(23)(24), this suggested a predominant macropinocytosis mechanism for its cell penetration (17). However, it was likely that such a predominant reliance on macropinocytosis was also conferred by the cargo type transported (streptavidin in that report).…”

“…Since these pioneering studies, MCa or analogues thereof proved powerful vectors for the cell entry of proteins, peptides (11), nanoparticles, or drugs such as doxorubicin (12)(13)(14)(15). Although the mode of cell penetration of MCa may vary according to cargo nature, cell type, or chemical linkage employed, the data gathered so far suggest that the peptide may enter cells according to two priming steps onto the plasma membrane: first an interaction with proteoglycans with an affinity in the micromolar range, followed by a second interaction with negatively charged lipids which occurs with greater affinity (16,17). The mode of cell entry of MCa is not altered by the absence of proteoglycans, but simply reduced quantitatively, suggesting that proteoglycans do not orient the mode of cell penetration.…”

d-Maurocalcine, a Pharmacologically Inert Efficient Cell-penetrating Peptide Analogue

“…Amiloride inhibited cell entry of FAM-D-MCa by 53.4 Ϯ 22%, which is less than when streptavidine is used as cargo. Interestingly, methyl-␤-cyclodextrin, which inhibits lipid raft-dependent endocytosis, also inhibits mildly the cell penetration of FAM-D-MCa (by 15.8 Ϯ 27%), which was not observed for streptavidine as cargo (14). This result indicates that macropinocytosis remains the major mode of FAM-D-MCa cell penetration by endocytosis.…”

“…These ratios between both distributions differ from the one observed using fluorescent streptavidine as cargo. With streptavidine, we have shown that the macropinocytosis inhibitor, amiloride, blocks 80% of cell entry (14). Here, we quantified by fluorescence-activated cell sorting (FACS) the effects of nocodazole (microtubule formation inhibitor), (Fig.…”

“…At the structural level, the peptide folds along an inhibitor cystine knot motif with three disulfide bridges connected according to the pattern Cys 3 -Cys 17 , Cys 10 -Cys 21 , and Cys 16 -Cys 32 . Landmark properties of CPP are also found in L-MCa because (i) 12 of the 33 residues of L-MCa are positively charged, (ii) many of the charged residues are critical for cell penetration, and (iii) L-MCa interacts with critical membrane components required for cell penetration, such as proteoglycans and negatively charged lipids (13,14). Altogether, these considerations made L-MCa the first example of a folded and oxidized peptide toxin acting as CPP.…”