title: Carbohydrate binding and resistance to proteolysis control insecticidal activity of Griffonia simplicifolia lectin II.
abstract: Griffonia simplicifolia leaf lectin II (GSII), a plant defense protein against certain insects, consists of an N-acetylglucosamine (GlcNAc)-binding large subunit with a small subunit having sequence homology to class III chitinases. Much of the insecticidal activity of GSII is attributable to the large lectin subunit, because bacterially expressed recombinant large subunit (rGSII) inhibited growth and development of the cowpea bruchid, Callosobruchus maculatus (F). Site-specific mutations were introduced into rGSII to generate proteins with altered GlcNAc binding, and the different rGSII proteins were evaluated for insecticidal activity when added to the diet of the cowpea bruchid. At pH 5.5, close to the physiological pH of the cowpea bruchid midgut lumen, rGSII recombinant proteins were categorized as having high (rGSII, rGSII-Y134F, and rGSII-N196D mutant proteins), low (rGSII-N136D), or no (rGSII-D88N, rGSII-Y134G, rGSII-Y134D, and rGSII-N136Q) GlcNAc-binding activity. Insecticidal activity of the recombinant proteins correlated with their GlcNAc-binding activity. Furthermore, insecticidal activity correlated with the resistance to proteolytic degradation by cowpea bruchid midgut extracts and with GlcNAc-specific binding to the insect digestive tract. Together, these results establish that insecticidal activity of GSII is functionally linked to carbohydrate binding, presumably to the midgut epithelium or the peritrophic matrix, and to biochemical stability of the protein to digestive proteolysis.
authors.affiliation: Department of Entomology, Purdue University, West Lafayette, IN 47907, USA.
authors.forename: K
authors.initials: K
authors.lastname: Zhu-Salzman
authors.forename: R E
authors.initials: RE
authors.lastname: Shade
authors.forename: H
authors.initials: H
authors.lastname: Koiwa
authors.forename: R A
authors.initials: RA
authors.lastname: Salzman
authors.forename: M
authors.initials: M
authors.lastname: Narasimhan
authors.forename: R A
authors.initials: RA
authors.lastname: Bressan
authors.forename: P M
authors.initials: PM
authors.lastname: Hasegawa
authors.forename: L L
authors.initials: LL
authors.lastname: Murdock
mesh_terms: D001665:Binding Sites; D050260:Carbohydrate Metabolism; D007306:Insecticides / Q000378:metabolism / Q000494:pharmacology*; D037102:Lectins / Q000378:metabolism / Q000494:pharmacology*; D037121:Plant Lectins; D010940:Plant Proteins / Q000378:metabolism / Q000494:pharmacology*; D010944:Plants / Q000378:metabolism*
chemical_list: D007306:Insecticides; D037102:Lectins; D037121:Plant Lectins; D010940:Plant Proteins