Enhanced conversion of plant biomass into glucose using transgenic rice-produced endoglucanase for cellulosic ethanol

Abstract

The catalytic domain of Acidothermus cellulolyticus thermostable endoglucanase gene (encoding for endo-1,4-beta-glucanase enzyme or E1) was constitutively expressed in rice. Molecular analyses of T1 plants confirmed presence and expression of the transgene. The amount of E1 enzyme accounted for up to 4.9% of the plant total soluble proteins, and its accumulation had no apparent deleterious effects on plant growth and development. Approximately 22 and 30% of the cellulose of the Ammonia Fiber Explosion (AFEX)-pretreated rice and maize biomass respectively was converted into glucose using rice E1 heterologous enzyme. As rice is the major food crop of the world with minimal use for its straw, our results suggest a successful strategy for producing biologically active hydrolysis enzymes in rice to help generate alcohol fuel, by substituting the wasteful and polluting practice of rice straw burning with an environmentally friendly technology.

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

“…Thermophilic cell wall-degrading enzymes with optimal temperatures over 50°C can be expressed in plants without disrupting cell wall integrity because they tend to perform poorly in normal plant growth conditions [76]. Hydrolases from thermophilic microorganisms such as Acidothermus cellulolyticus and Thermomonospora fusca, which are able to hydrolyze cellulose at high temperatures (81 and 75°C respectively), have been expressed in various plant species with no harmful effects [37,56,[77][78][79], while still having the desired effect of improving the ease of breakdown of plant material after harvest. Moreover, thermophilic enzymes are activated during post-harvest processing providing more advantages and applications in cellulosic ethanol production including simplifying processing and reducing exogenous enzyme loading [38].…”

“…Feedstocks with established genetic modification technology such as maize, rice, sugarcane, and poplar have been successfully transformed to express cell wall-degrading enzymes [55,77,79,80]. Several examples of expression of cell walldegrading enzymes in lignocellulosic biomass feedstock are provided in Table 1.…”

“…For example, plant produced recombinant glycosyl hydrolases can hydrolyze natural cellulosic biomass such as corn stover and rice straw [69,78]. Furthermore, several studies have shown that protein extracted from glycosyl hydrolase expressing transgenic plants can supplement exogenous enzymes during enzymatic hydrolysis [56,77,78]. Cellobiohydrolases expressed in corn were shown to enhance the performance of commercial cellulases in the degradation of pretreated sugarcane bagasse [83].…”

“…The cost of producing, extracting, and recovering rice-derived endoglucanases is currently estimated to be $9.64/kg enzymes ($0.64/gal) under the condition that 5 % TSP of endoglucanases is produced from rice straw [77]. Although this estimated cost is still far higher than the target price ($0.1/ gal), the situation could be improved by increasing expression of enzymes in planta and decreasing cost of enzyme extraction and recovery.…”

Genetically Engineered Crops for Biofuel Production: Regulatory Perspectives

“…Sufficient crop residue will need to remain on the soil to maintain soil quality. The E1 endoglucanase was the first enzyme engineered to be expressed in all tissues of corn (Ransom et al, 2007) and rice (Oraby et al, 2007) plants. Similar to the E1 tobacco, there was no visible phenotype in these transgenic plants compared to wild type lines yet the E1 was active on CMC.…”