Microbiology and Biotechnology, University of Hamburg, 22609, Hamburg, Germany.
Institute of Catalysis, Consejo Superior de Investigaciones Científicas, 28049, Madrid, Spain.
Sci Rep. 2018 Sep 14;8(1):13823. doi: 10.1038/s41598-018-32059-0.
Biotin is an essential cofactor required for carboxylation and decarboxylation reactions in all domains of life. While biotin biosynthesis in most Bacteria and Eukarya is well studied, the complete pathway for this vitamer in Archaea is still not known. Detailed genome searches indicated the presence of possible bio gene clusters only in Methanococcales and Thaumarchaeota. Therefore, we analysed the functionality of the predicted genes bioA, bioB, bioD and bioF in the Thaumarchaeon Nitrososphaera gargensis Ga2.9 which are essential for the later steps of biotin synthesis. In complementation tests, the gene cluster-encoded N. gargensis bioABD genes except bioF restored growth of corresponding E. coli Rosetta-gami 2 (DE3) deletion mutants. To find out how biotin biosynthesis is initiated, we searched the genome for a possible bioH analogue encoding a pimeloyl-ACP-methylester carboxylesterase. The respective amino acid sequence of the ORF estN1 showed weak conserved domain similarity to this class of enzymes (e-value 3.70e). Remarkably, EstN1 is a promiscuous carboxylesterase that complements E. coli ΔbioH and Mesorhizobium loti ΔbioZ mutants for growth on biotin-free minimal medium. Additional 3D-structural models support the hypothesis that EstN1 is a BioH analogue. Thus, this is the first report providing experimental evidence that Archaea carry functional bio genes.
生物素是所有生命领域中羧化和脱羧反应所需的必需辅酶。虽然大多数细菌和真核生物的生物素生物合成研究得很好，但古菌中这种维生素的完整途径仍不清楚。详细的基因组搜索表明，只有甲烷球菌目和奇古菌门中存在可能的生物基因簇。因此，我们分析了产甲烷菌 Nitrososphaera gargensis Ga2.9 中预测的生物素合成后期所必需的基因 bioA、bioB、bioD 和 bioF 的功能。在互补测试中，除了 bioF 之外，基因簇编码的 N. gargensis bioABD 基因恢复了相应的大肠杆菌 Rosetta-gami 2 (DE3) 缺失突变体的生长。为了找出生物素生物合成是如何开始的，我们在基因组中搜索了可能编码 pimeloyl-ACP-甲酯羧基酯酶的 bioH 类似物。ORF estN1 的相应氨基酸序列与该酶类具有较弱的保守结构域相似性（e 值 3.70e）。值得注意的是，EstN1 是一种混杂的羧基酯酶，可补充大肠杆菌ΔbioH 和 Mesorhizobium loti ΔbioZ 突变体在不含生物素的基础培养基上的生长。额外的 3D 结构模型支持 EstN1 是 BioH 类似物的假设。因此，这是第一个提供实验证据表明古菌携带功能性生物基因的报告。
