Zhao Shuting, Deng Dongtao, Wan Tianzheng, Feng Jie, Deng Lei, Tian Qianyi, Wang Jiayu, Aiman Umm E, Mukhaddi Balym, Hu Xiaofeng, Chen Shaolin, Qiu Ling, Huang Lili, Wei Yahong
State Key Laboratory of Crop Stress Biology for Arid Areas, College of Life Sciences, Biomass Energy Center for Arid and Semi-Arid Lands, Northwest A&F University, Yangling, Shaanxi, 712100, People's Republic of China.
Vrije University Amsterdam, De Boelelaan 1105, 1081 HV, Amsterdam, Netherlands.
Biotechnol Biofuels Bioprod. 2024 Feb 15;17(1):25. doi: 10.1186/s13068-024-02470-z.
Bioconversion of plant biomass into biofuels and bio-products produces large amounts of lignin. The aromatic biopolymers need to be degraded before being converted into value-added bio-products. Microbes can be environment-friendly and efficiently degrade lignin. Compared to fungi, bacteria have some advantages in lignin degradation, including broad tolerance to pH, temperature, and oxygen and the toolkit for genetic manipulation.
Our previous study isolated a novel ligninolytic bacterial strain Erwinia billingiae QL-Z3. Under optimized conditions, its rate of lignin degradation was 25.24% at 1.5 g/L lignin as the sole carbon source. Whole genome sequencing revealed 4556 genes in the genome of QL-Z3. Among 4428 protein-coding genes are 139 CAZyme genes, including 54 glycoside hydrolase (GH) and 16 auxiliary activity (AA) genes. In addition, 74 genes encoding extracellular enzymes are potentially involved in lignin degradation. Real-time PCR quantification demonstrated that the expression of potential ligninolytic genes were significantly induced by lignin. 8 knock-out mutants and complementary strains were constructed. Disruption of the gene for ELAC_205 (laccase) as well as EDYP_48 (Dyp-type peroxidase), ESOD_1236 (superoxide dismutase), EDIO_858 (dioxygenase), EMON_3330 (monooxygenase), or EMCAT_3587 (manganese catalase) significantly reduced the lignin-degrading activity of QL-Z3 by 47-69%. Heterologously expressed and purified enzymes further confirmed their role in lignin degradation. Fourier transform infrared spectroscopy (FTIR) results indicated that the lignin structure was damaged, the benzene ring structure and groups of macromolecules were opened, and the chemical bond was broken under the action of six enzymes encoded by genes. The abundant enzymatic metabolic products by EDYP_48, ELAC_205 and ESOD_1236 were systematically analyzed via liquid chromatography-mass spectrometry (LC-MS) analysis, and then provide a speculative pathway for lignin biodegradation. Finally, The activities of ligninolytic enzymes from fermentation supernatant, namely, LiP, MnP and Lac were 367.50 U/L, 839.50 U/L, and 219.00 U/L by orthogonal optimization.
Our findings provide that QL-Z3 and its enzymes have the potential for industrial application and hold great promise for the bioconversion of lignin into bioproducts in lignin valorization.
将植物生物质转化为生物燃料和生物产品会产生大量木质素。这些芳香族生物聚合物在转化为高附加值生物产品之前需要被降解。微生物能够以环境友好的方式高效降解木质素。与真菌相比，细菌在木质素降解方面具有一些优势，包括对pH、温度和氧气的广泛耐受性以及遗传操作工具。
我们之前的研究分离出了一种新型木质素降解细菌菌株欧文氏菌QL-Z3。在优化条件下，以1.5 g/L木质素作为唯一碳源时，其木质素降解率为25.24%。全基因组测序显示QL-Z3基因组中有4556个基因。在4428个蛋白质编码基因中，有139个碳水化合物活性酶（CAZyme）基因，包括54个糖苷水解酶（GH）基因和16个辅助活性（AA）基因。此外，74个编码细胞外酶的基因可能参与木质素降解。实时荧光定量PCR分析表明，潜在的木质素降解基因的表达受到木质素的显著诱导。构建了8个基因敲除突变体和互补菌株。敲除ELAC_205（漆酶）、EDYP_48（Dyp型过氧化物酶）、ESOD_1236（超氧化物歧化酶）、EDIO_858（双加氧酶）、EMON_3330（单加氧酶）或EMCAT_3587（锰过氧化氢酶）基因显著降低了QL-Z3的木质素降解活性，降幅为47%-69%。异源表达和纯化的酶进一步证实了它们在木质素降解中的作用。傅里叶变换红外光谱（FTIR）结果表明，在由这些基因编码的六种酶的作用下，木质素结构被破坏，苯环结构和大分子基团被打开，化学键断裂。通过液相色谱-质谱联用（LC-MS）分析系统地分析了EDYP_48、ELAC_205和ESOD_1236产生的丰富酶促代谢产物，进而为木质素生物降解提供了一个推测途径。最后，通过正交优化，发酵上清液中木质素降解酶，即木质素过氧化物酶（LiP）、锰过氧化物酶（MnP）和漆酶（Lac）的活性分别为367.50 U/L、839.50 U/L和219.00 U/L。
我们的研究结果表明，QL-Z3及其酶具有工业应用潜力，在木质素增值中将木质素转化为生物产品方面具有广阔前景。
