Abstract
BackgroundStarch 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.ResultsA novel raw starch-digesting glucoamylase PoGA15A with high enzymatic activity was purified from Penicillium oxalicum GXU20 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 in Pichia 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.ConclusionsA novel raw starch-digesting glucoamylase from P. 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.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0636-5) contains supplementary material, which is available to authorized users.
Highlights
Starch is a very abundant and renewable carbohydrate and is an important feedstock for industrial applications
Purification and identification of a novel raw starch‐digesting enzyme from P. oxalicum GXU20 During enzyme purification, the ability to hydrolyze raw starch was monitored when raw cassava starch was used as a substrate
Glucose remained the sole hydrolysis product and its concentration gradually increased. These results clearly demonstrated that the purified enzyme was an exo-acting amylase that cleaves glycosidic bonds successively to release glucose from the non-reducing ends of starch chains, thereby indicating that the purified amylase from P. oxalicum GXU20 was a glucoamylase
Summary
Starch is a very abundant and renewable carbohydrate and is an important feedstock for industrial applications. Starch is one of the most abundant renewable carbohydrate reserves in a large variety of higher plants, such as cereals, tuberous plants, and legumes. It is a polymer of glucose, and mainly consists of amylose and amylopectin. Starch is the second most important and abundant source of carbon and energy in plants, and has large market demand and many applications in industry. Starch is an important feedstock in the fermentation industry and is widely saccharified and fermented to produce ethanol, which can be used as a basis for beverages or as an alternative biofuel [3]
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