title: Efficient hydrolysis of raw starch and ethanol fermentation: a novel raw starch-digesting glucoamylase fromPenicillium oxalicum.
abstract: BACKGROUND
  Starch is a very abundant and renewable carbohydrate and is an important feedstock for industrial applications. The conventional starch liquefaction and saccharification processes are energy-intensive, complicated, and not environmentally friendly. Raw starch-digesting glucoamylases are capable of directly hydrolyzing raw starch to glucose at low temperatures, which significantly simplifies processing and reduces the cost of producing starch-based products.
  
  
  RESULTS
  A novel raw starch-digesting glucoamylase PoGA15A with high enzymatic activity was purified fromPenicillium oxalicumGXU20 and biochemically characterized. The PoGA15A enzyme had a molecular weight of 75.4 kDa, and was most active at pH 4.5 and 65 °C. The enzyme showed remarkably broad pH stability (pH 2.0-10.5) and substrate specificity, and was able to degrade various types of raw starches at 40 °C. Its adsorption ability for different raw starches was consistent with its degrading capacities for the corresponding substrate. The cDNA encoding the enzyme was cloned and heterologously expressed inPichia pastoris. The recombinant enzyme could quickly and efficiently hydrolyze different concentrations of raw corn and cassava flours (50, 100, and 150 g/L) with the addition of α-amylase at 40 °C. Furthermore, when used in the simultaneous saccharification and fermentation of 150 g/L raw flours to ethanol with the addition of α-amylase, the ethanol yield reached 61.0 g/L with a high fermentation efficiency of 95.1 % after 48 h when raw corn flour was used as the substrate. An ethanol yield of 57.0 g/L and 93.5 % of fermentation efficiency were achieved with raw cassava flour after 36 h. In addition, the starch-binding domain deletion analysis revealed that SBD plays a very important role in raw starch hydrolysis by the enzyme PoGA15A.
  
  
  CONCLUSIONS
  A novel raw starch-digesting glucoamylase fromP. oxalicum, with high enzymatic activity, was biochemically, molecularly, and genetically identified. Its efficient hydrolysis of raw starches and its high efficiency during the direct conversion of raw corn and cassava flours via simultaneous saccharification and fermentation to ethanol suggests that the enzyme has a number of potential applications in industrial starch processing and starch-based ethanol production.
authors: State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources, Guangxi Key Laboratory of Subtropical Bioresources Conservation and Utilization, Key Laboratory of Ministry of Education for Microbial and Plant Genetic Engineering, College of Life Science and Technology, Guangxi University, 100 Daxue Road, Nanning, 530004 Guangxi People's Republic of China.
authors: Qiang-Sheng
authors: QS
authors: Xu
authors: State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources, Guangxi Key Laboratory of Subtropical Bioresources Conservation and Utilization, Key Laboratory of Ministry of Education for Microbial and Plant Genetic Engineering, College of Life Science and Technology, Guangxi University, 100 Daxue Road, Nanning, 530004 Guangxi People's Republic of China.
authors: Yu-Si
authors: YS
authors: Yan
authors: State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources, Guangxi Key Laboratory of Subtropical Bioresources Conservation and Utilization, Key Laboratory of Ministry of Education for Microbial and Plant Genetic Engineering, College of Life Science and Technology, Guangxi University, 100 Daxue Road, Nanning, 530004 Guangxi People's Republic of China.
authors: Jia-Xun
authors: JX
authors: Feng
keywords: Cassava starch; Corn starch; Ethanol; Gene cloning and expression; Penicillium oxalicum; Raw starch hydrolysis; Raw starch-digesting glucoamylase; Simultaneous saccharification and fermentation