■β-葡萄糖苷酶是纤维素降解过程中关键的限速酶,促进纤维二糖和纤维寡糖水解成葡萄糖。然而,许多β-葡萄糖苷酶的广泛应用受到其有限的热稳定性和低葡萄糖耐量的阻碍,特别是在高温和高葡萄糖环境中。
■本研究对属于GH1家族的β-葡萄糖苷酶基因进行了分析,表示为lqbg8,它是从位于腾冲的荷花温泉宏基因组库中分离出来的,中国。随后,该基因被克隆并在大肠杆菌BL21(DE3)中异源表达。post表达式,重组β-葡萄糖苷酶(LQBG8)通过Ni亲和色谱柱纯化,从而能够对其酶学性质进行深入的探索。
■LQBG8具有70°C的最佳温度和5.6的最佳pH。LQBG8在65°C和70°C孵育2小时后保留了其最大活性的100%和70%,分别。此外,即使在暴露于3.0-10.0的pH范围24小时后,LQBG8保留了大约80%的初始活性。值得注意的是,LQBG8的酶促能力在葡萄糖浓度高达3M时保持显著,保留超过60%的相对活性。以纤维二糖为底物对LQBG8的动力学参数进行了表征,Km和Vmax值为28±1.9mg/mL和55±3.2μmol/min/mg,分别。此外,将LQBG8(浓度为0.03mg/mL)引入常规纤维素酶反应系统后,在24小时内玉米秸秆的葡萄糖产量提高了43.7%。分子动力学模拟为LQBG8的嗜热性质提供了有价值的见解,将其稳健的稳定性归因于波动的减少,构象变化,与嗜温β-葡萄糖苷酶相比,结构刚性提高。
■求和,它的嗜热,热稳定,和葡萄糖耐受属性,使LQBG8成熟,可用于涵盖食品的不同领域的潜在应用,饲料,和木质纤维素乙醇的生产。
UNASSIGNED: β-Glucosidase serves as the pivotal rate-limiting enzyme in the cellulose degradation process, facilitating the hydrolysis of cellobiose and cellooligosaccharides into glucose. However, the widespread application of numerous β-glucosidases is hindered by their limited thermostability and low glucose tolerance, particularly in elevated-temperature and high-glucose environments.
UNASSIGNED: This study presents an analysis of a β-glucosidase gene belonging to the GH1 family, denoted lqbg8, which was isolated from the metagenomic repository of Hehua hot spring located in Tengchong,
China. Subsequently, the gene was cloned and heterologously expressed in Escherichia coli BL21(DE3). Post expression, the recombinant β-glucosidase (LQBG8) underwent purification through a Ni affinity chromatography column, thereby enabling the in-depth exploration of its enzymatic properties.
UNASSIGNED: LQBG8 had an optimal temperature of 70°C and an optimum pH of 5.6. LQBG8 retained 100 and 70% of its maximum activity after 2-h incubation periods at 65°C and 70°C, respectively. Moreover, even following exposure to pH ranges of 3.0-10.0 for 24 h, LQBG8 retained approximately 80% of its initial activity. Notably, the enzymatic prowess of LQBG8 remained substantial at glucose concentrations of up to 3 M, with a retention of over 60% relative activity. The kinetic parameters of LQBG8 were characterized using cellobiose as substrate, with Km and Vmax values of 28 ± 1.9 mg/mL and 55 ± 3.2 μmol/min/mg, respectively. Furthermore, the introduction of LQBG8 (at a concentration of 0.03 mg/mL) into a conventional cellulase reaction system led to an impressive 43.7% augmentation in glucose yield from corn stover over a 24-h period. Molecular dynamics simulations offered valuable insights into LQBG8\'s thermophilic nature, attributing its robust stability to reduced fluctuations, conformational changes, and heightened structural rigidity in comparison to mesophilic β-glucosidases.
UNASSIGNED: In summation, its thermophilic, thermostable, and glucose-tolerant attributes, render LQBG8 ripe for potential applications across diverse domains encompassing food, feed, and the production of lignocellulosic ethanol.