Manipulating corn germplasm to increase recombinant protein accumulation

Abstract

Summary Using plants as biofactories for industrial enzymes is a developing technology. The application of this technology to plant biomass conversion for biofuels and biobased products has potential for significantly lowering the cost of these products because of lower enzyme production costs. However, the concentration of the enzymes in plant tissue must be high to realize this goal. We describe the enhancement of the accumulation of cellulases in transgenic maize seed as a part of the process to lower the cost of these dominant enzymes for the bioconversion process. We have used breeding to move these genes into elite and high oil germplasm to enhance protein accumulation in grain. We have also explored processing of the grain to isolate the germ, which preferentially contains the enzymes, to further enhance recovery of enzyme on a dry weight basis of raw materials. The enzymes are active on microcrystalline cellulose to release glucose and cellobiose.

Expression of Glycosyl Hydrolases in Lignocellulosic Feedstock: An Alternative for Affordable Cellulosic Ethanol Production

“…Acidothermus cellulolyticus endoglucanases were accumulated to over 16 % seed TSP in maize seed [29]. These levels of expression have also been shown to be stable over generations [34]. Despite these successes, in order for the expression of enzymes in planta to be effective, significantly higher expression levels are required or the enzymes must be produced more universally throughout the plant to increase per plant enzyme production.…”

“…These include studies showing expression levels up to 26 % total soluble protein (TSP) in Arabidopsis and 50 % TSP in tobacco leaves [29][30][31][32][33][34][35][36][37]. Although there is potential for high-level enzyme production in these plant systems, the expression of cell wall-degrading enzymes in the biomass of the lignocellulosic feedstocks themselves has added advantages [38].…”

Plant Molecular Farming: Much More than Medicines

“…Plant cell suspension cultures can be improved by high-throughput screening to identify the most productive cells and use them to produce high-yielding monoclonal cell lines (26). In contrast, the yields in whole plants can be increased by breeding and selection among the best performing primary transformants, which identifies those with stable transgenes at permissive integration sites and pairs them with the optimal genetic background (27,28). Process optimization strategies are similarly diverse.…”

“…The scale of production provides the greatest benefit compared to fermenter-based systems because it costs much more to scale up fermenter infrastructure than to grow additional hectares of a production crop, even if the crop is grown in greenhouses rather than open fields (108). We consider here the development of transgenic maize lines expressing cellobiohydrolase (CBH1) as a case study (115)(116)(117).…”

“…This process takes 4 years if a winter nursery is used to double the generations achieved per year but is worth the effort because large increases in yield are achieved. In the case of the CBH1 lines, the production hybrids achieved a twentyfold increase in yield over the original T1 seeds (116). During the breeding process, excess grain was used to optimize extraction and purification methods for product characterization and to test downstream applications in biomass conversion (117).…”