Ascertaining the biochemical function of an essential pectin methylesterase in the gut microbe Bacteroides thetaiotaomicron.
Ascertaining the biochemical function of an essential pectin methylesterase in the gut microbe Bacteroides thetaiotaomicron.
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DOI：
10.1074/jbc.ra120.014974
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发表时间：
2020-12-25
期刊：
影响因子：
--
通讯作者：
Ndeh D
中科院分区：
文献类型：
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作者：
Duan CJ;Baslé A;Liberato MV;Gray J;Nepogodiev SA;Field RA;Juge N;Ndeh D
果胶是人类肠道菌群的主要膳食营养来源。著名的肠道微生物拟杆菌（Bacteroides thetaiotaomicron）最近被证明编码一个新的果胶甲基酯酶家族的创始成员（BT1017），该家族对复杂果胶鼠李糖半乳糖醛酸- ii （RG-II）的代谢至关重要。然而，该家族的生化和结构知识缺乏。在这里，我们发现BT1017对rg - ii衍生的低聚糖ΔBT1017oligoB的代谢至关重要，该低聚糖是由BT1017缺失突变体（ΔBT1017）在苹果汁碳水化合物提取物生长过程中产生的。利用酶、质谱和核磁共振方法对ΔBT1017oligoB进行结构分析，发现它是一种双甲基化的非低聚糖(GlcA-β1,4-(2- o - me - xyl -α1,3)- fuc -α1,4-(GalA-β1,3)- rha -α1,3- api -β1,2-(Araf-α1,3)-(GalA-α1,4)-GalA)，含有RG-II主链及其侧链的组分。我们发现BT1017的催化模块采用α/β-水解酶折叠，由中心扭曲的10链β-片夹在几个α-螺旋中组成。这构成了果胶甲基酯酶的新折叠，主要是右手性β-螺旋蛋白。生物信息学分析显示，该家族的主要序列来自人类肠道微生物群的突出属，包括拟杆菌和普雷沃氏菌。我们的研究结果不仅突出了该酶家族在果胶代谢中的关键作用，而且为B. thetaiotaommicron适应人类肠道的分子基础提供了新的见解。
Pectins are a major dietary nutrient source for the human gut microbiota. The prominent gut microbe Bacteroides thetaiotaomicron was recently shown to encode the founding member (BT1017) of a new family of pectin methylesterases essential for the metabolism of the complex pectin rhamnogalacturonan-II (RG-II). However, biochemical and structural knowledge of this family is lacking. Here, we showed that BT1017 is critical for the metabolism of an RG-II–derived oligosaccharide ΔBT1017oligoB generated by a BT1017 deletion mutant (ΔBT1017) during growth on carbohydrate extract from apple juice. Structural analyses of ΔBT1017oligoB using a combination of enzymatic, mass spectrometric, and NMR approaches revealed that it is a bimethylated nonaoligosaccharide (GlcA-β1,4-(2-O-Me-Xyl-α1,3)-Fuc-α1,4-(GalA-β1,3)-Rha-α1,3-Api-β1,2-(Araf-α1,3)-(GalA-α1,4)-GalA) containing components of the RG-II backbone and its side chains. We showed that the catalytic module of BT1017 adopts an α/β-hydrolase fold, consisting of a central twisted 10-stranded β-sheet sandwiched by several α-helices. This constitutes a new fold for pectin methylesterases, which are predominantly right-handed β-helical proteins. Bioinformatic analyses revealed that the family is dominated by sequences from prominent genera of the human gut microbiota, including Bacteroides and Prevotella. Our re-sults not only highlight the critical role played by this family of enzymes in pectin metabolism but also provide new insights into the molecular basis of the adaptation of B. thetaiotaomicron to the human gut.