title: Structure and glycolipid binding properties of the nematicidal protein Cry5B.
abstract: Crystal (Cry) proteins are globally used in agriculture as proteinaceous insecticides. They have also been recently recognized to have great potential as anthelmintic agents in targeting parasitic roundworms (e.g., hookworms). The most extensively characterized of the anthelmintic Cry proteins is Cry5B. We report here the 2.3 Å resolution structure of the proteolytically activated form of Cry5B. This structure, which is the first for a nematicidal Cry protein, shows the familiar three-domain arrangement seen in insecticidal Cry proteins. However, domain II is unusual in that it more closely resembles a banana lectin than it does other Cry proteins. This result is consistent with the fact that the receptor for Cry5B consists of a set of invertebrate-specific glycans (attached to lipids) and also suggests that domain II is important for receptor binding. We found that not only galactose but also N-acetylgalactosamine (GalNAc) is an efficient competitor for binding between Cry5B and glycolipids. GalNAc is one of the core arthroseries tetrasaccharides of the Cry5B receptor and galactose an antennary sugar that emanates from this core. These and prior data suggest that the minimal binding determinant for Cry5B consists of a core GalNAc and two antennary galactoses. Lastly, the protoxin form of Cry5B was found to bind nematode glycolipids with a specificity equal to that of activated Cry5B, but with lower affinity. This suggests that the initial binding of Cry5B protoxin to glycolipids can be stabilized at the nematode cell surface by proteolysis. These results lay the groundwork for the design of effective Cry5B-based anthelmintics.
authors.affiliation: Department of Chemistry and Biochemistry, 9500 Gilman Drive, University of California, San Diego, La Jolla, CA 92093-0375, USA.
authors.forename: Fan
authors.initials: F
authors.lastname: Hui
authors.forename: Ulrike
authors.initials: U
authors.lastname: Scheib
authors.forename: Yan
authors.initials: Y
authors.lastname: Hu
authors.forename: Ralf J
authors.initials: RJ
authors.lastname: Sommer
authors.forename: Raffi V
authors.initials: RV
authors.lastname: Aroian
authors.forename: Partho
authors.initials: P
authors.lastname: Ghosh
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D003460:Crystallization; D004731:Endotoxins / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D006017:Glycolipids / Q000378:metabolism*; D006460:Hemolysin Proteins / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009348:Nematoda / Q000187:drug effects*; D011485:Protein Binding; D017386:Sequence Homology, Amino Acid
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006017:Glycolipids; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis