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Discovery and Expression of Thermostable LPMOs from Thermophilic Fungi for Producing Efficient Lignocellulolytic Enzyme Cocktails.
Applied Biochemistry and Biotechnology ( IF 3.1 ) Pub Date : 2019-12-02 , DOI: 10.1007/s12010-019-03198-5
Dhruv Agrawal 1 , Neha Basotra 1 , Venkatesh Balan 2 , Adrian Tsang 3 , Bhupinder Singh Chadha 1
Affiliation  

In this study, two novel thermostable lytic polysaccharide monooxygenases (LPMOs) were cloned from thermophilic fungus Scytalidium thermophilum (PMO9D_SCYTH) and Malbranchea cinnamomea (PMO9D_MALCI) and expressed in the methylotrophic yeast Pichia pastoris X33. The purified PMO9D_SCYTH was active at 60 °C (t1/2 = 60.58 h, pH 7.0), whereas, PMO9D_MALCI was optimally active at 50 °C (t1/2 = 144 h, pH 7.0). The respective catalytic efficiency (kcat/Km) of PMO9D_SCYTH and PMO9D_MALCI determined against avicel in presence of H2O2 was (6.58 × 10-3 and 1.79 × 10-3 mg-1 ml min-1) and carboxy-methylcellulose (CMC) (1.52 × 10-1 and 2.62 × 10-2 mg-1 ml min-1). The HRMS analysis of products obtained after hydrolysis of avicel and CMC showed the presence of both C1 and C4 oxidized oligosaccharides, in addition to phylogenetic tree constructed with other characterized type 1 and 3 LPMOs demonstrated that both LPMOs belongs to type-3 family of AA9s. The release of sugars during saccharification of acid/alkali pretreated sugarcane bagasse and rice straw was enhanced upon replacing one part of commercial enzyme Cellic CTec2 with these LPMOs.

中文翻译:

从嗜热真菌中发现并表达热稳定的LPMO,以生产高效的木质纤维素分解酶鸡尾酒。

在这项研究中,从嗜热真菌Scytalidium thermophilum(PMO9D_SCYTH)和Malbranchea cinnamomea(PMO9D_MALCI)中克隆了两种新型的热稳定性裂解多糖单加氧酶(LPMO),并在甲基营养酵母巴斯德毕赤酵母X33中表达。纯化的PMO9D_SCYTH在60°C(t 1/2 = 60.58 h,pH 7.0)下具有活性,而PMO9D_MALCI在50°C(t 1/2 = 144 h,pH 7.0)上具有最佳活性。在H 2 O 2存在下,PMO9D_SCYTH和PMO9D_MALCI对avicel的催化效率(k cat / K m)为(6.58×10 -3和1.79×10-3 mg -1 ml min -1)和羧甲基纤维素(CMC)(1.52×10 -1和2.62×10 -2 mg -1 ml min -1)。avicel和CMC水解后获得的产物的HRMS分析表明,除了用其他特征性1型和3型LPMO构建的系统树以外,C 1和C 4氧化寡糖均存在,这表明这两个LPMO均属于CLP的3型家族。 AA9s。用这些LPMO代替商业酶Cellic CTec2的一部分,可以增强酸/碱预处理的甘蔗渣和稻草糖化过程中糖的释放。
更新日期:2019-12-11
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