Zhao Fang, Yu Chun-Mei, Sun Hai-Ning, Zhao Long-Sheng, Ding Hai-Tao, Cao Hai-Yan, Chen Yin, Qin Qi-Long, Zhang Yu-Zhong, Li Ping-Yi, Chen Xiu-Lan
Marine Biotechnology Research Center, State Key Laboratory of Microbial Technology, Shandong University, Qingdao, China; MOE Key Laboratory of Evolution and Marine Biodiversity, Frontiers Science Center for Deep Ocean Multispheres and Earth System & College of Marine Life Sciences, Ocean University of China, Qingdao, China; Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao, China.
Marine Biotechnology Research Center, State Key Laboratory of Microbial Technology, Shandong University, Qingdao, China.
J Biol Chem. 2023 Sep;299(9):105116. doi: 10.1016/j.jbc.2023.105116. Epub 2023 Jul 29.
Xylans are polysaccharides composed of xylose and include β1,4-xylan, β1,3-xylan, and β1,3/1,4-mixed-linkage xylan (MLX). MLX is widely present in marine red algae and constitutes a significant organic carbon in the ocean. Xylanases are hydrolase enzymes that play an important role in xylan degradation. While a variety of β1,4-xylanases and β1,3-xylanases involved in the degradation of β1,4-xylan and β1,3-xylan have been reported, no specific enzyme has yet been identified that degrades MLX. Herein, we report the characterization of a new MLX-specific xylanase from the marine bacterium Polaribacter sp. Q13 which utilizes MLX for growth. The bacterium secretes xylanases to degrade MLX, among which is Xyn26A, an MLX-specific xylanase that shows low sequence similarities (<27%) to β1,3-xylanases in the glycoside hydrolase family 26 (GH26). We show that Xyn26A attacks MLX precisely at β1,4-linkages, following a β1,3-linkage toward the reducing end. We confirm that Xyn26A and its homologs have the same specificity and mode of action on MLX, and thus represent a new xylanase group which we term as MLXases. We further solved the structure of a representative MLXase, AlXyn26A. Structural and biochemical analyses revealed that the specificity of MLXases depends critically on a precisely positioned β1,3-linkage at the -2/-1 subsite. Compared to the GH26 β1,3-xylanases, we found MLXases have evolved a tunnel-shaped cavity that is fine-tuned to specifically recognize and hydrolyze MLX. Overall, this study offers a foremost insight into MLXases, shedding light on the biochemical mechanism of bacterial degradation of MLX.
木聚糖是由木糖组成的多糖，包括β1,4-木聚糖、β1,3-木聚糖和β1,3/1,4-混合连接木聚糖（MLX）。MLX广泛存在于海洋红藻中，是海洋中重要的有机碳成分。木聚糖酶是在木聚糖降解过程中发挥重要作用的水解酶。虽然已经报道了多种参与β1,4-木聚糖和β1,3-木聚糖降解的β1,4-木聚糖酶和β1,3-木聚糖酶，但尚未鉴定出能降解MLX的特定酶。在此，我们报道了从海洋细菌极地杆菌属Q13中鉴定出一种新的MLX特异性木聚糖酶的特性，该细菌利用MLX进行生长。该细菌分泌木聚糖酶来降解MLX，其中Xyn26A是一种MLX特异性木聚糖酶，与糖苷水解酶家族26（GH26）中的β1,3-木聚糖酶具有较低的序列相似性（<27%）。我们发现Xyn26A精确地在β1,4-连接键处攻击MLX，沿着β1,3-连接键向还原端进行。我们证实Xyn26A及其同源物对MLX具有相同的特异性和作用模式，因此代表了一个新的木聚糖酶组，我们将其称为MLX酶。我们进一步解析了一种代表性MLX酶AlXyn26A的结构。结构和生化分析表明，MLX酶的特异性关键取决于-2/-1亚位点处精确位置的β1,3-连接键。与GH26β1,3-木聚糖酶相比，我们发现MLX酶进化出了一个隧道形腔，该腔经过精细调整以特异性识别和水解MLX。总体而言，这项研究为MLX酶提供了最前沿的见解，揭示了细菌降解MLX的生化机制。
