Currently, the sole strategy for managing food hypersensitivity involves strict avoidance of the trigger. Several alternate strategies for the treatment of food allergies are currently under study. Also being explored is the process of eliminating allergenic proteins from crop plants. Legumes are a rich source of protein and are an essential component of the human diet. Unfortunately, legumes, including soybean and peanut, are also common sources of food allergens. Four protein families and superfamilies account for the majority of legume allergens, which include storage proteins of seeds (cupins and prolamins), profilins, and the larger group of pathogenesis-related proteins. Two strategies have been used to produce hypoallergenic legume crops: (1) germplasm lines are screened for the absence or reduced content of specific allergenic proteins and (2) genetic transformation is used to silence native genes encoding allergenic proteins. Both approaches have been successful in producing cultivars of soybeans and peanuts with reduced allergenic proteins. However, it is unknown whether the cultivars are actually hypoallergenic to those with sensitivity. This review describes efforts to produce hypoallergenic cultivars of soybean and peanut and discusses the challenges that need to be overcome before such products could be available in the marketplace.

Soybean is a common allergenic food; thus, a comprehensive characterization of all the proteins that cause allergy is crucial to the development of effective diagnostic and immunotherapeutic strategies. A cDNA library was constructed from seven stages of developing soybean seeds to investigate candidate allergens. We searched the library for cDNAs encoding a seed-specific biotinylated protein (SBP) based on its allergenicity in boiled lentils. A full-length cDNA clone was retrieved and expressed as a 75.6-kDa His-tagged recombinant protein (rSBP) in Escherichia coli. Western immunoblotting of boiled bacterial extracts demonstrated specific IgE binding to rSBP, which was further purified by metal affinity and anion exchange chromatographies. Of the 23 allergic sera screened by ELISA, 12 contained IgEs specific to the purified rSBP. Circular dichroism spectroscopy revealed a predominantly unordered structure consistent with SBP's heat stability. The natural homologues (nSBP) were the main proteins isolated from soybean and peanut embryos after streptavidin affinity purification, yet they remained low-abundance proteins in the seed as confirmed by LC-MS/MS. Using capture ELISAs, the soybean and peanut nSBPs were bound by IgEs in 78 and 87% of the allergic sera tested. The soybean nSBP was purified to homogeneity and treatments with different denaturing agents before immunoblotting highlighted the diversity of its IgE epitopes. In vitro activation of basophils was assessed by flow cytometry in a cohort of peanut-allergic children sensitized to soybean. Stronger and more frequent (38%) activations were induced by nSBP-soy compared to the major soybean allergen, Gly m 5. SBPs may represent a novel class of biologically active legume allergens with the structural resilience to withstand many food-manufacturing processes.

High-level expression of transgenes is essential for cost-effective production of valuable pharmaceutical proteins in plants. However, transgenic proteins often accumulate in plants at low levels. Low levels of protein accumulation can be caused by many factors including post-transcriptional gene silencing (PTGS) and/or rapid turnover of the transgenic proteins. We have developed an Amplicon-plus Targeting Technology (APTT), by using novel combination of known techniques that appears to overcome both of these factors. By using this technology, we have successfully expressed the highly-labile L1 protein of canine oral papillomavirus (COPV L1) by infecting transgenic tobacco plants expressing a suppressor of post-transcriptional gene silencing (PTGS) with a PVX amplicon carrying a gene encoding L1, and targeting the vaccine protein into the chloroplasts. Further, a scalable ''wound-and-agrospray'' inoculation method has been developed that will permit high-throughput Agrobacterium inoculation of Nicotiana tabacum, and a spray-only method (named ''agrospray'') for use with N. benthamiana to allow large-scale application of this technology. The good yield and short interval from inoculation to harvest characteristic of APTT, combined with the potential for high-throughput achieved by use of the agrospray inoculation protocol, make this system a very promising technology for producing high value recombinant proteins, especially those known to be highly labile, in plants for a wide range of applications including producing vaccines against rapidly evolving pathogens and for the rapid response needed to meet bio-defense emergencies.

Due to an unfortunate misunderstanding, the co-corresponding author and the electronic supplementary materials were not properly identified in the original publication. The correct representation of the authors and their affiliations is listed below.The online version of the original article can be found at http:// dx

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