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.

Laboratory study of the effects of leek lectin (APA) in transgenic tobacco plants on the development of cotton leafworm Spodoptera littoralis (Lepidoptera: Noctuidae)

“…Additional support for such a mode of action comes from the observation that for the Griffonia simplicifolia lectin (GS-II) there is a clear correlation between receptor binding and toxicity towards the cowpea bruchid Callosobruchus maculatus (Zhu-Salzman et al, 1998). In addition, lectins may also act as a feeding deterrent leading to reduce food consumption.…”

Insecticide Activity of Lectins and Secondary Metabolites

“…The precise mechanisms of insecticidal action of lectins remain unknown, though it has been suggested that this entomotoxic activity seems to depend upon the carbohydrate recognition property they exhibit. Plant lectins with affinity for N-acetylglucosamine and chitin-binding property are able to bind chitin and glycosylated proteins of the peritrophic matrix, interfering in the digestion and absorption of nutrients (Tellam et al, 1999;Peumans and Van Damme, 1995;Zhu-Salzman et al, 1998;Zhu-Salzman and Salzman, 2001;Carlini and Grossi-de-Sá, 2002;Macedo et al, 2004;Macedo et al, 2007). The peritrophic matrix constitutes a membrane found in the midgut that separates the contents of the gut lumen from the digestive epithelial cells.…”

“…Lectins have deleterious effects against larvae, developing stages and mature forms of insects from orders Coleoptera, Diptera, Hemiptera, Homoptera, Hymenoptera, Isoptera, Lepidoptera and Neuroptera (Murdock et al, 1990;Eisemann et al, 1994;Powell et al, 1995;Zhu-Salzman et al, 1998;Bandyopadhyay et al, 2001;Isidro et al, 2001;Hogervorst et al, 2006;Kaur et al, 2006;Coelho et al, 2007;Macedo et al, 2007;Fitches et al, 2008;Sá et al, 2008;Coelho et al, 2009;Sá et al, 2009c;Silva et al, 2009;Napoleão et al, 2011;Souza et al, 2011b). Insecticide activity of lectin is generally evaluated by bioassays that incorporate the lectin into artificial diets offered to insects, with insects dying from nutritional deprivation.…”

Plant‐insect interactions: what can we learn from plant lectins?

“…If a lectin is resistant to proteolytic digestion by the insect, it can also bind to the insect gut. Hence, resistance to proteolysis is another aspect of the toxicity of a lectin, as shown by Zhu-Salzman et al (1998), Zhu-Salzman and Salzman (2001), and Lagarda-Diaz et al (2009). Foissac et al (2000 indicate that the binding of a lectin is not the only parameter for its toxicity level.…”