Conserved active site cysteine residue of archaeal THI4 homolog is essential for thiamine biosynthesis in Haloferax volcanii
Conserved active site cysteine residue of archaeal THI4 homolog is essential for thiamine biosynthesis in Haloferax volcanii
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DOI：
10.1186/s12866-014-0260-0
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发表时间：
2014-10-28
期刊：
影响因子：
4.2
通讯作者：
Maupin-Furlow, Julie A.
中科院分区：
文献类型：
--
作者：
Hwang, Sungmin;Cordova, Bryan;Maupin-Furlow, Julie A.
背景：硫胺素（维生素B1）是由某些酵母、真菌、植物、原生动物、细菌和古细菌从头合成的。对古细菌合成硫胺素的途径，特别是硫接力形成噻唑环的途径了解甚少。古细菌含有细菌（ThiS-ThiF）和真核生物（THI4）蛋白的结构同源物，这些蛋白通过不同的机制将硫动员到噻唑环前体。结果：基于比较基因组分析，预测嗜盐古菌通过细菌途径合成硫胺素的嘧啶部分，初步表明细菌的ThiS- thif型机制也用于合成噻唑环，其中硫载体ThiS首先被thif催化的腺苷化激活。火山卤铁（Haloferax volcanii, UbaA）唯一的ThiF同源物被删除，但在缺乏硫胺素的情况下，这对生长没有影响。使用真核THI4型硫传递最初被认为不太可能用于古生菌的硫胺素生物合成，因为酵母THI4p的活性位点半胱氨酸残基通过自杀机制将硫提供给噻唑环，在许多古生菌的THI4同源物中被组氨酸残基取代，这些被描述为d -核糖-1,5-二磷酸异构酶。包括Hfx在内的嗜盐古菌的THI4同源物。火山菌（HVO_0665, HvThi4）的半胱氨酸残基（Cys165）与酵母THI4p的保守活性位点半胱氨酸（Cys205）相对应，与产甲烷菌和热球菌的半胱氨酸残基不同。HVO_0665的缺失产生了硫胺素缺陷，该缺陷被HVO_0665的野生型拷贝反式补充，但没有编码HvThi4 C165A变体的修饰基因。结论：根据我们的研究结果，我们认为古菌Hfx。volcanii利用酵母thi4型机制进行硫接力，形成硫胺素的噻唑环。我们将这一发现扩展到一个相对较大的古菌群，包括盐古菌，铵氧化古菌，以及一些甲烷菌和焦球菌物种，通过观察这些生物编码具有保守活性位点半胱氨酸残基的THI4同源物，这可能用于硫胺素的生物合成。因此，具有保守半胱氨酸活性位点的IPR002922 THI4家族古细菌成员应该重新研究其在硫胺素生物合成中的作用。
Background: Thiamine (vitamin B1) is synthesized de novo by certain yeast, fungi, plants, protozoans, bacteria and archaea. The pathway of thiamine biosynthesis by archaea is poorly understood, particularly the route of sulfur relay to form the thiazole ring. Archaea harbor structural homologs of both the bacterial (ThiS-ThiF) and eukaryotic (THI4) proteins that mobilize sulfur to thiazole ring precursors by distinct mechanisms.Results: Based on comparative genome analysis, halophilic archaea are predicted to synthesize the pyrimidine moiety of thiamine by the bacterial pathway, initially suggesting that also a bacterial ThiS-ThiF type mechanism for synthesis of the thiazole ring is used in which the sulfur carrier ThiS is first activated by ThiF-catalyzed adenylation. The only ThiF homolog of Haloferax volcanii (UbaA) was deleted but this had no effect on growth in the absence of thiamine. Usage of the eukaryotic THI4-type sulfur relay was initially considered less likely for thiamine biosynthesis in archaea, since the active-site cysteine residue of yeast THI4p that donates the sulfur to the thiazole ring by a suicide mechanism is replaced by a histidine residue in many archaeal THI4 homologs and these are described as D-ribose-1,5-bisphosphate isomerases. The THI4 homolog of the halophilic archaea, including Hfx. volcanii (HVO_0665, HvThi4) was found to differ from that of methanogens and thermococci by having a cysteine residue (Cys165) corresponding to the conserved active site cysteine of yeast THI4p (Cys205). Deletion of HVO_0665 generated a thiamine auxotroph that was trans-complemented by a wild-type copy of HVO_0665, but not the modified gene encoding an HvThi4 C165A variant.Conclusions: Based on our results, we conclude that the archaeon Hfx. volcanii uses a yeast THI4-type mechanism for sulfur relay to form the thiazole ring of thiamine. We extend this finding to a relatively large group of archaea, including haloarchaea, ammonium oxidizing archaea, and some methanogen and Pyrococcus species, by observing that these organisms code for THI4 homologs that have a conserved active site cysteine residue which is likely used in thiamine biosynthesis. Thus, archaeal members of IPR002922 THI4 family that have a conserved cysteine active site should be reexamined for a function in thiamine biosynthesis.