Lin Hui, Hildebrand Amanda, Kasuga Takao, Fan Zhiliang
Biological and Agricultural Engineering Department, University of California, Davis, One Shields Avenue, Davis, CA 95616, United States.
Department of Plant Pathology, University of California, Davis, One Shields Avenue, Davis, CA 95616, United States; United States Department of Agriculture-Agricultural Research Service, Davis, CA 95616, United States.
Enzyme Microb Technol. 2017 Apr;99:25-31. doi: 10.1016/j.enzmictec.2016.12.009. Epub 2017 Jan 6.
In a cellulosic biorefinery, the cellulase enzymes needed for hydrolysis are one of the major contributors to high processing costs, while the hydrolysis product, cellobiose, has strong inhibition to the cellulases. In this study, we report engineering recombinant Neurospora crassa strains which are able to produce cellobionate, an organic acid, from cellulose without any enzyme addition. Recombinant strains were constructed by heterologously expressing laccase genes from different sources under different promoters in N. crassa F5Δmus-51Δace-1Δcre-1ΔndvB which has six out of seven β-glucosidase (bgl), two transcription factor (cre1 and ace-1), and the cellobionate phosphorylase (ndvB) genes deleted. The strain expressing laccase from Botrytis aclada under a copper metallothionein promoter (HL10) produced the highest laccase activity. N. crassa HL10 produced 47.4mM cellobionate from cellulose without any enzyme addition. The yield of cellobionate from hydrolyzed cellulose was about 94.5%. Conversion of cellobiose to cellobionate improved cellulose conversion and increases product yield.
在纤维素生物精炼厂中，水解所需的纤维素酶是加工成本高昂的主要因素之一，而水解产物纤维二糖对纤维素酶具有强烈的抑制作用。在本研究中，我们报道了构建重组粗糙脉孢菌菌株，该菌株能够在不添加任何酶的情况下从纤维素产生有机酸盐纤维二糖酸。通过在粗糙脉孢菌F5Δmus-51Δace-1Δcre-1ΔndvB中不同启动子下异源表达来自不同来源的漆酶基因构建重组菌株，该菌株已缺失七个β-葡萄糖苷酶（bgl）中的六个、两个转录因子（cre1和ace-1）以及纤维二糖酸磷酸化酶（ndvB）基因。在铜金属硫蛋白启动子（HL10）下表达来自灰葡萄孢漆酶的菌株产生了最高的漆酶活性。粗糙脉孢菌HL10在不添加任何酶的情况下从纤维素产生了47.4 mM的纤维二糖酸。水解纤维素生成纤维二糖酸的产率约为94.5%。纤维二糖向纤维二糖酸的转化提高了纤维素转化率并增加了产物产率。
