Yin Yi-Rui, Sang Peng, Xian Wen-Dong, Li Xin, Jiao Jian-Yu, Liu Lan, Hozzein Wael N, Xiao Min, Li Wen-Jun
State Key Laboratory of Biocontrol and Guangdong Provincial Key Laboratory of Plant Resources, School of Life Sciences, Sun Yat-sen University, Guangzhou, China.
College of Agriculture and Biological Science, Dali University, Dali, China.
Front Microbiol. 2018 Dec 18;9:3149. doi: 10.3389/fmicb.2018.03149. eCollection 2018.
The bioconversion of lignocellulose in various industrial processes, such as biofuel production, requires the degradation of cellulose. YIM 77502 is an aerobic, Gram-positive actinomycete that can efficiently degrade crystalline cellulose by extracellular cellulases. Genomic analysis of identified 9 cellulase and 11 β-glucosidase genes that could potentially encode proteins that digest cellulose. Extracellular proteome characterization of cell-free culture supernatant by liquid chromatography tandem mass spectrometry analysis revealed that 4 of these cellulases and 2 of these β-glucosidases functioned during cellulose hydrolysis. Thin-layer chromatography analysis revealed extracellular β-glucosidases play a major role in carboxyl methyl cellulose (CMC) degradation of products in culture supernatants. In this study, 2 of the identified secreted β-glucosidases, AaBGL1 and AaBGL2, were functionally expressed in and found to have β-glucosidase activity with wide substrate specificities, including for -nitrophenyl β-D-glucopyranoside (pNPG), -nitrophenyl-beta-D-cellobioside (pNPC), and cellobiose. Moreover, AaBGL1 and AaBGL2 had high tolerances for glucose. After adding these β-glucosidases to commercial cellulases, the degradation rates of CMC, Avicel, birch sawdust, and corncob powder increased by 37, 42, 33, and 9%, respectively. Overall, this work identifies an alternative potential source of β-glucosidases with potential applications in commercial cellulose utilization and the bioenergy industry.
在各种工业过程（如生物燃料生产）中，木质纤维素的生物转化需要纤维素的降解。YIM 77502是一种好氧革兰氏阳性放线菌，能够通过细胞外纤维素酶高效降解结晶纤维素。对其进行基因组分析，鉴定出9个纤维素酶基因和11个β-葡萄糖苷酶基因，这些基因可能编码消化纤维素的蛋白质。通过液相色谱串联质谱分析对其无细胞培养上清液进行细胞外蛋白质组表征，结果显示其中4种纤维素酶和2种β-葡萄糖苷酶在纤维素水解过程中发挥作用。薄层色谱分析表明，细胞外β-葡萄糖苷酶在培养上清液中羧甲基纤维素（CMC）降解产物过程中起主要作用。在本研究中，鉴定出的2种分泌型β-葡萄糖苷酶AaBGL1和AaBGL2在大肠杆菌中实现了功能表达，并且发现它们对包括对硝基苯基β-D-吡喃葡萄糖苷（pNPG）、对硝基苯基-β-D-纤维二糖苷（pNPC）和纤维二糖在内的多种底物具有广泛的β-葡萄糖苷酶活性。此外，AaBGL1和AaBGL2对葡萄糖具有高耐受性。将这些β-葡萄糖苷酶添加到商业纤维素酶中后，CMC、微晶纤维素、桦木屑和玉米芯粉的降解率分别提高了37%、42%、33%和9%。总体而言，这项工作确定了一种β-葡萄糖苷酶的潜在替代来源，其在商业纤维素利用和生物能源行业具有潜在应用价值。
