Bioactive molecules from ciliates: Structure, activity, and applicative potential

Abstract

Ciliates are a rich source of molecules synthesized to socialize, compete ecologically, and interact with prey and predators. Their isolation from laboratory cultures is often straightforward, permitting the study of their mechanisms of action and their assessment for applied research. This review focuses on three classes of these bioactive molecules: (i) water-borne, cysteine-rich proteins that are used as signaling pheromones in self/nonself recognition phenomena; (ii) cell membrane-associated lipophilic terpenoids that are used in interspecies competitions for habitat colonization; (iii) cortical granule-associated molecules of various chemical nature that primarily serve offence/defense functions.

Keywords: climacostol; euplotins; pheromones; resorcinol; signaling proteins; terpenoids.

FIGURE 1

Amino acid sequences and NMR solution structures of Euplotes pheromones. (A) For each species‐specific pheromone family, represented by the members listed between brackets, only the consensus sequence is reported. The conserved cysteines are highlighted in red, and X stands for any amino acid. The sequence of the E. crassus pheromone Ec‐α, the only one known from a sub‐family distinct from that which includes the other pheromone sequences, is reported individually and written in the single letter code. Analogously, the sequences of the E. octocarinatus pheromone Phr4 and E. raikovi pheromone Er‐23 are reported individually, as they represent structurally deviant family members. The molecular structures of E. raikovi pheromones Er‐1 (PDB code, 1ERC) and Er‐23 (PDB code, 1HA8), E. nobilii pheromone En‐6 (PDB code, 2JMS), and E. petzi pheromone Ep‐1 (PDB code, 2N2S) are shown (in ribbon diagrams) as representative of each pheromone family. The disulfide bonds stabilizing the molecular structures are shown as yellow ball‐and‐stick diagrams. The helix labeled h3 in the Er‐1 pheromone structure is highlighted in red to indicate the tight intraspecific and interspecific conservation of the structural backbone. N and C identify the molecule amino and carboxyl termini, respectively. (B) Very simplified version of the Euplotes phylogenetic tree, articulated into six clades (boxes), showing the positions of the five species analyzed for the pheromone structures

FIGURE 2

Crystal structures of E. raikovi pheromones Er‐1 and Er‐13. (A) Amino acid sequences and molecular structures of the two pheromones. In the amino acid sequences, the three helical segments (h1, h2 and h3) are boxed, and the six cysteine residues are indicated by progressive Roman numerals and connected according to their disulfide pairings represented by yellow ball‐and‐stick diagrams. Helix 3 is highlighted in red to indicate the central role played in establishing the contacts that pack molecules into crystals. (B) Perspective view (on the left) and top view (on the right) of the one‐dimensional linear chains, propagating along a two‐fold screw‐symmetry rotation axis (red symbol), that Er‐1 and Er‐13 molecules equally form in the crystal taking rigorously alternating up and down orientations, as indicated by the positions of the amino (N) and carboxyl (C) termini. The Er‐1 and Er‐13 chains include a common protein–protein contact interface (green bars), that is equally derived from burying amino acid side‐chains mostly residing on h3. However, they differ in the geometrical arrangement of the molecules, as denoted by observing that Er‐1 molecules expose both h1 and h2 parallel to the chain sides, while Er‐13 molecules expose only h2. In the Er‐1 crystal (PDBs, 1ERL, 2ERL and 6E6O), this difference reflects in a further side‐by‐side chain association into bidimensional layers (not shown) and in two‐fold symmetries specific of the space group C2. In the Er‐13 crystal (PDB, 6E6N), it reflects in a mutual arrangement of the chains at 90° between one another (not shown), which prevents the layer formation and is consistent with four‐fold screw symmetries distinctive of the space group P4₃

FIGURE 3

The four distinct classes of chemical structures of lipophilic terpenoids, designated euplotins, characterized from different marine species of Euplotes. Arrows indicate biosynthetic derivatives from precursor forms

FIGURE 4

Bioactive molecules of different chemical nature from cortical granules of species of Pseudokeronopsis, Spirostomum and Climacostomum. In the three lactone molecules, different side chains attached to a same carbon skeleton are indicated by R₁ and R₂. In the three synthetic derivatives (indicated by arrows and named AN1, AN2 and MOMO) of native climacostol, the side chain modifications are in bold