The ACEII recombinant Trichoderma reesei QM9414 strains with enhanced xylanase production and its applications in production of xylitol from tree barks

Abstract

BackgroundACEII transcription factor plays a significant role in regulating the expression of cellulase and hemicellulase encoding genes. Apart from ACEII, transcription factors such as XYR1, CRE1, HAP2/3/5 complex and ACEI function in a coordinated pattern for regulating the gene expression of cellulases and hemicellulases. Studies have demonstrated that ACEII gene deletion results in decreased total cellulase and xylanase activities with reduced transcript levels of lignocellulolytic enzymes.ResultsIn this study, we have successfully transformed the ACEII transcription factor encoding gene in Trichoderma reesei to significantly improve its degrading abilities. Transformation experiments on parental strain T. reesei QM9414 has resulted in five genetically engineered strains T/Ace2-2, T/Ace2-5, T/Ace2-8, T/Ace5-4 and T/Ace10-1. Among which, T/Ace2-2 has exhibited significant increase in enzyme activity by twofolds, when compared to parental strain. The T/Ace2-2 was cultured on growth substrates containing 2% bark supplemented with (a) sugar free + MA medium (b) glucose + MA medium and (c) xylose + MA medium. The bark degradation efficiency of genetically modified T/Ace2-2 strain was assessed by analyzing the xylitol production yield using HPAEC. By 6th day, about 10.52 g/l of xylitol was produced through enzymatic conversion of bark (2% bark + MA + xylose) by the T/Ace2-2 strain and by 7th day the conversion rate was found to be 0.21 g/g. Obtained results confirmed that bark growth medium supplemented with d-xylose has profoundly increased the conversion rate of bark by T/Ace2-2 strain when compared to sugar free and glucose supplemented growth media. Results obtained from scanning electron microscopy has endorsed our current results. Bark samples inoculated with T/Ace2-2 strain has showed large number of degraded cells with clearly visible cavities and fractures, by exposing the microfibrillar interwoven complex.ConclusionWe propose a cost effective and ecofriendly method for the degradation of lignocellulosic biomass such as bark to produce xylitol by using genetically modified T. reesei. Efficient conversion rate and production yield obtained in our current study provides a great scope for the xylitol industries, as our method bypasses the pretreatment of bark achieving clean and low-cost xylitol production.

Effects of the Transcription Factor Ace2 from Trichoderma reesei on Cellulase and Hemicellulase Expression in Trichoderma orientalis EU7-22

“…These results indicated that the expression of ace2 in T. orientalis largely improved the β-glucosidase activity and xylanase activity. In T. reesei, deletion of ace2 gene resulted in reduced expression levels of cellulases on Solka oc cellulose [18,19] and overexpression of ace2 gene exhibited signi cant increase in [20]. However, in contrast with the claim of Ace2 activating the cellulase production, most of the cellulase activities decreased after expressing ace2 in EU7-22.…”

“…Some relevant studies were given by Xiong et al They have successfully transformed the Ace2 transcription factor encoding gene in T. reesei and gotten ve genetically engineered strains. Among which, the result displayed that only one has exhibited a signi cant increase in enzyme activity, but the other four showed lower total cellulase activities than the parental strain T. reesei QM9414 [20]. Also, Ace2 was not subject to carbon catabolite repression, because in the △cre1 strain no increased expression levels of ace2 gene were observed.…”

Factors regulating cellulolytic gene expression in filamentous fungi: an overview

“…The role of Ace2 as a positive regulator of cellulase expression was further confirmed in another study where the overexpression of Ace2 in T. reesei led to a twofold increase in cellulase activity in the recombinant T/Ace2-2 strain when grown in a medium containing a mixture of hardwoods supplemented with glucose or xylose. [ 60 ].…”

Production and applications of xylanases – an overview

“…Perez-Gonzalez and coworkers (1996) Filamentous fungi are preferred expression systems because of the ease in handling fungal cultivation and strain improvement they have already undergone (Nevalainenm et al, 2005). Xylanases have been expressed in T. reesei, A. kawachii, and A. niger (Xiong et al, 2016;Qiu et al, 2016;Takahashi et al, 2013). P. pastoris has been explored for the expression of transgenic xylanase.…”