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Reducing Exogenous Enzyme Requirements in Biofuels Production By Metabolic Engineering

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Conventional corn dry grind ethanol process requires exogenous alpha and glucoamylases enzymes to breakdown starch into glucose, which is fermented to ethanol by yeast. Currently these alpha and glucoamylases enzymes are added externally in liquid form during the liquefaction and saccharification process respectively. This study investigates the use of genetically engineered corn and yeast, which can eliminate or minimize the use of these external enzymes and potentially reduce the operating cost and simplify the ethanol production process. The amylase corn used in the study has been genetically engineered to produce α-amylase in endosperm, which eliminates the need of external enzyme addition during liquefaction. Due to high expression levels of enzymes, only a small amount of the corn is mixed with the conventional dent corn. Similarly, the GA yeast investigated in this study has also been engineered to produce endogenous glucoamylases and provide novel metabolic pathways for high ethanol yields. The fermentation characteristics of amylase corn mix (15% amylase corn) using GA yeast (evaluated at 0% and 50% loadings of glucoamylases enzyme) were evaluated using laboratory dry grind process and the results were compared with the control treatments (100% dent corn, conventional yeast). The ethanol yield of amylase corn mix was 4.16% higher than that of dent corn using conventional yeast. For dent corn, no significant differences were observed in the ethanol yields between the control and using GA yeast without any addition of glucoamylases. Addition of only 50% glucoamylases along with GA yeast use resulted in a high ethanol concentration (18.96%) for amylase mixed corn. The ethanol yields for dent corn and amylase corn were found 3.08 and 3.04 L/kg corn respectively, which were 7.83 and 2.08% higher than that of conventional yeast at 100% glucoamylases loading. The findings from this study demonstrated the potential of getting high ethanol yields and reducing exogenous enzyme requirements using metabolic engineered corn and yeast.