Comparative insights into the saccharification potentials of a relatively unexplored but robust Penicillium funiculosum glycoside hydrolase 7 cellobiohydrolase
Comparative insights into the saccharification potentials of a relatively unexplored but robust Penicillium funiculosum glycoside hydrolase 7 cellobiohydrolase
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DOI：
10.1186/s13068-017-0752-x
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发表时间：
2017-03-20
影响因子：
6.3
通讯作者：
Yazdani, Syed Shams
中科院分区：
文献类型：
--
作者：
Ogunmolu, Funso Emmanuel;Jagadeesha, Navya Bhatt Kammachi;Yazdani, Syed Shams
背景：GH7纤维素生物水解酶（CBH1）对纤维素的分解至关重要。我们以前观察到该酶是高纤维素水解子囊菌-真菌青霉（NCIM1228）的活性纤维素水解分泌组中最显性的蛋白。为了了解其对纤维素生物质糖化的贡献，并将其与工业原料木霉（trichoderma reesei）的GH7纤维素生物水解酶进行比较，我们对真菌基因组中唯一鉴定并存在的GH7纤维素生物水解酶进行了天然纯化和功能表征。结果：两种酶的稳定性存在边际差异，P. funiculosum （PfCBH1）在pH 4.4下的最佳热中点（T-m）为68℃，而T. reesei （TrCBH1）在pH 4.7下的最佳热中点（T-m）为65℃。然而，与TrCBH1相比，PfCBH1对对硝基苯-a-d-乳吡喃苷（pNPL）的结合亲和力（k -m）低约3倍，周转率（k(cat)）高18倍，催化效率高6倍，酶抑制剂复合物平衡解离常数（k -i）高26倍。虽然这两种酶都能水解纤维素低聚物（G2-G6）和微晶纤维素，释放出纤维素二糖和葡萄糖作为主要产物，但PfCBH1的倾向更大。在相同条件下，我们同样观察到预处理小麦秸秆与其他核心纤维素酶串联水解时的这种趋势。以TrCBH1结构（PDB ID: 8CEL）为模板建立的同源模型进行分子动力学模拟，使我们能够直接研究底物和产物对蛋白质动力学的影响。虽然催化三元组- exdxxe基序-在两种酶之间是保守的，但在包围催化路径的区域中观察到细微的变化，并强调了与功能的关系。结论：据我们所知，这是第一篇关于高溶子囊菌- p中CBH1的全面和比较描述的报道。在同样的酶的背景下，从工业工作马t。reesei。我们的研究表明，PfCBH1是工业纤维素酶鸡尾酒中来自T. reesei的CBH1的可行替代品。
Background: GH7 cellobiohydrolases (CBH1) are vital for the breakdown of cellulose. We had previously observed the enzyme as the most dominant protein in the active cellulose-hydrolyzing secretome of the hypercellulolytic ascomycete- Penicillium funiculosum (NCIM1228). To understand its contributions to cellulosic biomass saccharification in comparison with GH7 cellobiohydrolase from the industrial workhorse-Trichoderma reesei, we natively purified and functionally characterized the only GH7 cellobiohydrolase identified and present in the genome of the fungus.Results: There were marginal differences observed in the stability of both enzymes, with P. funiculosum (PfCBH1) showing an optimal thermal midpoint (T-m) of 68 degrees C at pH 4.4 as against an optimal T-m of 65 degrees C at pH 4.7 for T. reesei (TrCBH1). Nevertheless, PfCBH1 had an approximate threefold lower binding affinity (K-m), an 18-fold higher turnover rate (k(cat)), a sixfold higher catalytic efficiency as well as a 26-fold higher enzyme-inhibitor complex equilibrium dissociation constant (K-i) than TrCBH1 on p-nitrophenyl-a-d-lactopyranoside (pNPL). Although both enzymes hydrolyzed cellooligomers (G2-G6) and microcrystalline cellulose, releasing cellobiose and glucose as the major products, the propensity was more with PfCBH1. We equally observed this trend during the hydrolysis of pretreated wheat straws in tandem with other core cellulases under the same conditions. Molecular dynamic simulations conducted on a homology model built using the TrCBH1 structure (PDB ID: 8CEL) as a template enabled us to directly examine the effects of substrate and products on the protein dynamics. While the catalytic triads-EXDXXE motifs-were conserved between the two enzymes, subtle variations in regions enclosing the catalytic path were observed, and relations to functionality highlighted.Conclusion: To the best of our knowledge, this is the first report about a comprehensive and comparative description of CBH1 from hypercellulolytic ascomycete-P. funiculosum NCIM1228, against the backdrop of the same enzyme from the industrial workhorse-T. reesei. Our study reveals PfCBH1 as a viable alternative for CBH1 from T. reesei in industrial cellulase cocktails.