Critical amino acid residues of maurocalcine involved in pharmacology, lipid interaction and cell penetration

Abstract

Maurocalcine (MCa) is a 33-amino acid residue peptide that was initially identified in the Tunisian scorpion Scorpio maurus palmatus. This peptide triggers interest for three main reasons. First, it helps unravelling the mechanistic basis of Ca(2+) mobilization from the sarcoplasmic reticulum because of its sequence homology with a calcium channel domain involved in excitation-contraction coupling. Second, it shows potent pharmacological properties because of its ability to activate the ryanodine receptor. Finally, it is of technological value because of its ability to carry cell-impermeable compounds across the plasma membrane. Herein, we characterized the molecular determinants that underlie the pharmacological and cell-penetrating properties of maurocalcine. We identify several key amino acid residues of the peptide that will help the design of cell-penetrating analogues devoid of pharmacological activity and cell toxicity. Close examination of the determinants underlying cell penetration of maurocalcine reveals that basic amino acid residues are required for an interaction with negatively charged lipids of the plasma membrane. Maurocalcine analogues that penetrate better have also stronger interaction with negatively charged lipids. Conversely, less effective analogues present a diminished ability to interact with these lipids. These findings will also help the design of still more potent cell penetrating analogues of maurocalcine.

Small Efficient Cell-penetrating Peptides Derived from Scorpion Toxin Maurocalcine

“…This may represent an inhibitory region because of the presence of Glu 12 and Asp 15 , two negatively charged residues. The finding that mutation of Glu 12 to Ala enhances cell penetration of both MCa F (18) and MCa UF (11) further supports this conclusion. MCa UF25-33 -C turned out to have CPP properties also, even though this sequence did not confer a competitive advantage to other MCa CPP analogues as shown in Fig.…”

“…First, all of the charged peptides (8 tested) demonstrated CPP properties. These peptides all had the K 22 R 23 R 24 sequence in common, a cluster of basic amino acid residues shown to contribute to the dose efficacy of cell penetration of MCa F in an earlier study (18). Interestingly, removing the last 8 C-terminal amino acids of MCa had little impact on the cell penetration properties (if one compares MCa UF14 -25 -C with MCa UF14 -33 -C).…”

“…This strategy preserved the disulfide bridges and the three-dimensional structure of the analogues. Overall, mutations affected more seriously the pharmacology of MCa than the cell penetration properties (18). Many of the amino acids involved in RyR1 binding and pharmacology were located within the cluster of basic amino acids that presented sequence homology with the L-type Ca v 1.1 channel.…”

Multiple actions of φ-LITX-Lw1a on ryanodine receptors reveal a functional link between scorpion DDH and ICK toxins

“…LITX also does not form pores in the membrane, as shown by the absence of current when applied to a membrane without RyRs. It was previously noted that the calcines have a high net positive charge (+7) and that the basic residues are important for membrane permeability (39). In comparison, LITX has a lower net change of +4, which may explain its inability to pass rapidly through membranes (Fig.…”

“…1) and channel mean open time (Table S1) It has not been determined whether LITX and the scorpion calcines share one or more common RyR binding sites; however, their similar modes of action suggest it is possible. Mutagenesis studies of the calcines IpTxA and maurocalcine have shown that numerous basic and acidic residues contribute to the activity of these toxins (39,44,45). A conserved Arg at position 24 in the calcines is of particular interest because it is essential for the induction of substates, but seems to have only a small involvement in full RyR openings (38,45).…”