Proteome of Soybean Seed Exudates Contains Plant Defense-Related Proteins Active against the Root-Knot Nematode Meloidogyne incognita.

Several studies have described the effects of seed exudates against microorganisms, but only few of them have investigated the proteins that have defensive activity particularly against nematode parasites. This study focused on the proteins released in the exudates of soybean seeds and evaluated their nematicidal properties against Meloidogyne incognita. A proteomic approach indicated the existence of 63 exuded proteins, including ß-1,3-glucanase, chitinase, lectin, trypsin inhibitor, and lipoxygenase, all of which are related to plant defense. The presence of some of these proteins was confirmed by their in vitro activity. The soybean exudates were able to reduce the hatching of nematode eggs and to cause 100% mortality of second-stage juveniles (J2). The pretreatment of J2 with these exudates resulted in a 90% reduction of the gall number in tobacco plants. These findings suggest that the exuded proteins are directly involved in plant defense against soil pathogens, including nematodes, during seed germination.

Identification of an alpha-amylase inhibitor from Pterodon pubescens with ability to inhibit cowpea weevil digestive enzymes.

Cowpea seeds (Vigna ungiculata) are widely cultivated by poor farmers in Latin America and Africa and are often severely damaged by the cowpea weevil Callosobruchus maculatus. A proteinaceous inhibitor of cowpea weevil digestive enzymes, PpAI, was purified from white sucupira seeds (Pterodon pubescens) and biochemically characterized in this study. Proteins were extracted from seeds and precipitated with ammonium sulfate at 100% saturation. This fraction was applied onto a Red-sepharose CL-6B column, and the retained peak showed 70% inhibitory activity toward larval C. maculatus digestive alpha-amylases. The retained peak was then purified using an analytical reversed-phase HPLC column. Purified PpAI showed 65% inhibitory activity against larval C. maculatus enzymes. Enzymatic assays also showed that the purified P. pubescens inhibitor was unable to reduce the activity of mammalian alpha-amylases, suggesting specificity toward insect enzymes. Moreover, artificial seeds containing PpAI were able to reduce larval weight by 36% and cause 55% mortality. Mass spectrometry and SDS-PAGE analyses indicated that PpAI showed a molecular mass of approximately 5.0 kDa. This alpha-amylase inhibitor, coming from a native Cerrado plant, could be used to construct a genetically engineered cowpea with enhanced resistance against weevil pests.