Structure of the polypeptide crotamine from the Brazilian rattlesnake Crotalus durissus terrificus

Abstract

The crystal structure of the myotoxic, cell-penetrating, basic polypeptide crotamine isolated from the venom of Crotalus durissus terrificus has been determined by single-wavelength anomalous dispersion techniques and refined at 1.7 Å resolution. The structure reveals distinct cationic and hydrophobic surface regions that are located on opposite sides of the molecule. This surface-charge distribution indicates its possible mode of interaction with negatively charged phospholipids and other molecular targets to account for its diverse pharmacological activities. Although the sequence identity between crotamine and human β-defensins is low, the three-dimensional structures of these functionally related peptides are similar. Since crotamine is a leading member of a large family of myotoxic peptides, its structure will provide a basis for the design of novel cell-penetrating molecules.

Keywords: crotamine; natural cell-penetrating polypeptides; snake venoms.

Figure 1

(a) Structure of crotamine. The N- and C-termini and the cysteine residues are labelled. (b) The highly hydrophobic residues in pink are located on one side of the molecule and the positively charged residues in blue are on the opposite side. (c) Space-filling presentation of crotamine, highlighting the well defined amphipathic surface region in blue and pink.

Figure 2

(a) Cartoon plot of the crotamine trimer. (b) Surface-charge distribution in two orientations.

Figure 3

(a) Sequence alignment of antimicrobial peptides. Conserved Cys residues are indicated in dark grey. Crotamine (PDB entry 4gv5), defensin-like peptide 2 (PDB entry 1d6b), heliomicin (PDB entry 1i2u), toxin III (PDB entry 1lqq), Eucommia antifungal peptide 2 (PDB entry 1p9z), charybdotoxin (PDB entry 2crd), human β-defensin 1 (PDB entry 1iju), human β-defensin 2 (PDB entry 1fd4), human β-defensin 3 (PDB entry 1kj6) and human α-defensin 1 (PDB entry 2pm1). (b) Superposition of crotamine (cartoon plot in light grey) with hβD-1 (PDB entry 1iju, pink), hβD-2 (PDB entry 1fd4, red) and hβD-3 (PDB entry 1kj6, cyan). The corresponding C^α r.m.s.d. values are 1.8, 1.8 and 2.6 Å, respectively.

Figure 4

Structure comparison of crotamine with homologous antimicrobial and antimicrobial-like peptides from different organisms in two orientations. Column 1, the γ-core domain is shown in orange. Column 2, hydropathy plot overlay: dark orange/hydrophobic; light orange/intermediate; blue/hydrophilic. Column 3, surface charge: blue, basic (Arg, Lys); red, acidic (Asp, Glu). Row A, crotamine from C. durissus terrificus venom (PDB entry 4gv5); row B, defensin-like peptide 2 (1d6b); row C, heliomicin (1i2u); row D, toxin III (1lqq); row E, Eucommia antifungal peptide 2 (1p9z); Row F, charybdotoxin (2crd); row G, human β-­defensin 1 (1iju); row H, human β-defensin 2 (1fd4); row I, human β-­defensin 3 (1kj6); row J, human α-­defensin 1 (2pm1).