FIG. 1.

Molecular and functional characterization of longicin. (A) cDNA and deduced amino acid sequences and charge density of longicin. Locations of secondary structure elements are shown in the form of cylinders (α-helixes) and an arrow (β-strand). Italics indicate a putative signal sequence, the triangle indicates the mature peptide, the box indicates P4 peptide, and a bold underline marks the polyadenylation signal. (B) Alignment of longicin and the scorpion ion channel blocker (SICB) from Sahara scorpion. Asterisks indicate conserved residues. (C) Coomassie blue-stained recombinant longicins. (D) Endogenous longicin in tick protein extract. Detection was performed by two-dimensional immunoblot analysis with anti-longicin antibody. (E) Longicin expression induced by blood feeding. An equal amount of protein (three ticks) was loaded in each lane. Detection was performed by immunoblot analysis with anti-longicin antibody. Pre, prefeeding. (F) Immunohistochemical localization of endogenous longicin. Note the strong staining of the midgut epithelium (high magnification). Abbreviations: cu, cuticle; mg, midgut; sg, salivary gland; tr, trachea. (G) Bactericidal activity of longicin. Bacterial cells were exposed to recombinant longicin or synthetic peptides for 2 h at 37°C, and then the incubation mixture was spread on TSB agar plates. The results shown are from four plates from different batches. (H) Fungal cells were exposed to recombinant longicin or P4 peptide for 2 h at 37°C, and then the incubation mixture was spread on YTB agar plates. (I) Morphological changes in P. pastoris induced by longicin.