An Enterotoxin-Like Binary Protein from Pseudomonas protegens with Potent Nematicidal Activity

• Published in: Applied and environmental microbiology Vol. 83; no. 19
• Main Authors: Wei, Jun-Zhi, Siehl, Daniel L, Hou, Zhenglin, Rosen, Barbara, Oral, Jarred, Taylor, Christopher G, Wu, Gusui
• Format: Journal Article
• Language: English
• Published: United States American Society for Microbiology 01.10.2017
• Subjects: Adenosine diphosphate; Amino Acid Sequence; Animals; Antinematodal Agents - metabolism; Antinematodal Agents - pharmacology; Bacteria; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bacterial Proteins - pharmacology; Base Sequence; Caenorhabditis elegans; Caenorhabditis elegans - drug effects; Caenorhabditis elegans - growth & development; Cholera; Cholera toxin; E coli; Enterotoxins - genetics; Enterotoxins - metabolism; Enterotoxins - pharmacology; Enzymology and Protein Engineering; Escherichia coli; Food sources; Fungi; Gram-negative bacteria; Heat-labile enterotoxin; Infections; Insects; Larvae; Molecular Sequence Data; Nematoda - drug effects; Nematoda - growth & development; Nematodes; Predation; Protein purification; Proteins; Pseudomonas; Pseudomonas - chemistry; Pseudomonas - genetics; Pseudomonas - metabolism; Pseudomonas protegens; Public health; Soil bacteria; Soil microorganisms; Soils; Toxins; Waterborne diseases; Pseudomonas protegens; enterotoxin; enterotoxins; nematode; binary toxin; anti-nematode protein
• ISSN: 0099-2240; 1098-5336
• DOI: 10.1128/AEM.00942-17

Soil microbes are a major food source for free-living soil nematodes. It is known that certain soil bacteria have evolved systems to combat predation. We identified the nematode-antagonistic strain 15G2 from screening of microbes. Through protein purification we identified a binary protein, designated Pp-ANP, which is responsible for the nematicidal activity. This binary protein inhibits growth and development by arresting larvae at the L1 stage and killing older-staged worms. The two subunits, Pp-ANP1a and Pp-ANP2a, are active when reconstituted from separate expression in The binary toxin also shows strong nematicidal activity against three other free-living nematodes ( , , and sp.), but we did not find any activity against insects and fungi under test conditions, indicating specificity for nematodes. Pp-ANP1a has no significant identity to any known proteins, while Pp-ANP2a shows ∼30% identity to heat-labile enterotoxin (LT) subunit A and cholera toxin (CT) subunit A. Protein modeling indicates that Pp-ANP2a is structurally similar to CT/LT and likely acts as an ADP-ribosyltransferase. Despite the similarity, Pp-ANP shows several characteristics distinct from CT/LT toxins. Our results indicate that Pp-ANP is a new enterotoxin-like binary toxin with potent and specific activity to nematodes. The potency and specificity of Pp-ANP suggest applications in controlling parasitic nematodes and open an avenue for further research on its mechanism of action and role in bacterium-nematode interaction. This study reports the discovery of a new enterotoxin-like binary protein, Pp-ANP, from a strain. Pp-ANP shows strong nematicidal activity against larvae and older-staged worms. It also shows strong activity on other free-living nematodes ( , , and sp.). The two subunits, Pp-ANP1a and Pp-ANP2a, can be expressed separately and reconstituted to form the active complex. Pp-ANP shows some distinct characteristics compared with other toxins, including enterotoxin and cholera toxin. The present study indicates that Pp-ANP is a novel binary toxin and that it has potential applications in controlling parasitic nematodes and in studying toxin-host interaction.