Persistence and ecological implications of maize- expressed transgenic endo-1,4-β-D-glucanase (E1) in agricultural soils

Plant expression of thermostable endoglucanase (E1) [pdb:1ECE] has been proposed for improved conversion of lignocellulose to ethanol for fuel production. Since E1 was isolated from hot springs and has thermostable properties, it may have increased stability when released to agricultural soils during post-harvest residue decomposition. Endoglucanases are a subclass of cellulases and act by cleaving β-1,4 glycosidic bonds in cellulose polymers at amorphous sites. In addition, they are important in carbon cycling and can be useful indicators of possible ecological perturbations in soil. It is estimated that where genetically-engineered maize expressing E1 is grown, it will add 17-120 g ha y of thermostable endoglucanase in postharvest residues to receiving soils. The persistence and reactivity of residual E1 is being investigated in soil microcosm studies for soils amended with bacterial and plant-solubilized E1 as compared to soil endogenous activity and to activity added from a mesostable form of endoglucanase (Aspergillus and Trichoderma spp.). An optimized analytical method involving a carboxymethyl cellulose (CMC) substrate and dinitrosalicylic acid (DNS) detection method effectively assayed endoglucanase activity in amended and unamended soils and was used for determining E1 persistence in representative soils. Effects on soil carbon mineralization were determined by comparing CO2 evolution from soils amended with E1 and Wild-type corn tissue. Extraction and recovery of the mesostable comparator, bacterial E1, and plant-soluble E1 shows near-complete dissipation of activity within a 24-hour period. Carbon mineralization studies indicate no significant difference between soils amended with either the transgenic E1 or Wild-type maize tissue. The combined result from persistence and CO2 evolution studies suggest that maize expressing up to 30 g Mg of E1 in stover (120 g ha environmental load) would not persist in representative soils or significantly impact carbon cycling.