Molecular basis and its application of formaldehyde-fixing reactions in bacteria and Archaea.

细菌和古细菌中甲醛固定反应的分子基础及其应用。

• 批准号: 15380061
• 负责人: KATO Nobuo
• 金额: $ 10.37万
• 依托单位: KYOTO UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15380061/
• 关键词: methylotroph; formaldehyde; Pyrococcus horikoshii; Bacillus subtilis; Mycobacterium gastri; methanogen; Hps; Phi; メタノール; ホルムアルデヒド固定反応; Mycobacterium gastri MB19

Hexulose-6-phosphate synthase(Hps) and Phosphohexuloisomerase(Phi), two key enzymes in the ribulose monophosphate pathway, has been recently found in a number of bacterial and archaeal strains. In this study, we studied on (i)the regulation of the expresseion of hps and phi gene in bacteria, and (ii)enzymological properties and their physiological roles of hps and phi in archaea.(i)Two regulatory proteins (RmpR from a methylotroph Mycobacterium gastri MB19 and HxlR from a non-methylotroph Bacillus subtilis) were produced in E.coli and their abilities to DNA were tested. HxlR bound to the upstream region of the hps/phi operon and was shown to act as a positive regulator of the formaldehyde-inducible gene expression.(ii)The hyperthermophilic archaeon Pyrococcus horikoshii had a fused protein consisting of Hps and Phi. The entire fusion protein and separate Hps and Phi polypeptides were produced in E.coli, purified and biochemically characterized. The fused protein was found in a particulate fraction. It was suggested that the thermostability of the Phi moiety of the fused protein resulted from fusion with Hps. Fusion of Hps and Phi increased the catalytic efficiency of the net reaction. On the other hand the methanogenic archaeon Methanosarcina mazei Goel had a fused protein consisting of Hps and formaldehyde activating enzyme (Fae). Hps-Fae fused protein produced in E.coli exhibited both Hps and Phi activities. From the catalytic properties of Hps-Phi and Hps-Fae proteins from two archeal strains, it was suggested that these proteins are involved not only in the formaldehyde fixation (forward reaction) but also in the generation of ribose 5-phosphate (reverse reaction).

6-磷酸己酮糖合酶（Hps）和磷酸己酮糖异构酶（Phi）是核酮糖单磷酸途径中的两种关键酶，最近在许多细菌和古细菌菌株中被发现。在本研究中，我们研究了（i）细菌中hps和phi基因表达的调节，以及（ii）古细菌中hps和phi的酶学特性及其生理作用。（i）产生了两种调节蛋白（来自甲基营养型胃分枝杆菌MB19的RmpR和来自非甲基营养型枯草芽孢杆菌的HxlR） 在大肠杆菌中，并测试了它们的 DNA 能力。 HxlR 与 hps/phi 操纵子的上游区域结合，并被证明充当甲醛诱导基因表达的正调节因子。(ii)超嗜热古菌 Pyrococcus horikoshii 具有由 Hps 和 Phi 组成的融合蛋白。整个融合蛋白以及单独的 Hps 和 Phi 多肽在大肠杆菌中产生、纯化并进行生化表征。在颗粒部分中发现了融合蛋白。表明融合蛋白的Phi部分的热稳定性是由于与Hps融合所致。 Hps 和 Phi 的融合提高了净反应的催化效率。另一方面，产甲烷古菌 Methanosarcina mazei Goel 具有由 Hps 和甲醛激活酶 (Fae) 组成的融合蛋白。大肠杆菌中产生的 Hps-Fae 融合蛋白表现出 Hps 和 Phi 活性。从两个古菌菌株的 Hps-Phi 和 Hps-Fae 蛋白的催化特性来看，这些蛋白不仅参与甲醛固定（正向反应），而且还参与核糖 5-磷酸（逆向反应）的生成。

项目成果

HxlR, a member of the DUF24 protein family, is a DNA-binding protein that acts as a positive regulator of the formaldehyde-inducible hxLAB operon in Bacillus subtilis

HxlR 是 DUF24 蛋白家族的成员，是一种 DNA 结合蛋白，在枯草芽孢杆菌中充当甲醛诱导型 hxLAB 操纵子的正调节因子

• 发表时间: 2005
• 期刊: Mol.Microbiol. (印刷中)
• 作者: Yurimoto;H.;et al.
• 通讯作者: et al.

微生物の新規な代謝機能の解明とその応用に関する研究

微生物新代谢功能阐明及其应用研究

• 发表时间: 2004
• 期刊: 日本農芸化学会誌 78
• 作者: ^*Tateishi Y;^*Kawabe Y(^*co-first author);Chiba T;Murata S;Ichikawa K;Murayama A;Tanaka K;Baba T;Kato S;Yanagisawa J;加藤暢夫
• 通讯作者: 加藤暢夫

Mitsui, R: "Formaldehyde fixation contributes to detoxification for growth of a nonmethylotrophic Burkholderia cepacia TM1"Appl.Environ.Microbiol.. 69. 6128-6132 (2003)

Mitsui, R：“甲醛固定有助于非甲基营养性洋葱伯克霍尔德菌 TM1 生长的解毒”Appl.Environ.Microbiol.. 69. 6128-6132 (2003)

メチロトローフのホルムアルデヒド対策

甲基营养菌中的甲醛防治措施

• 发表时间: 2003
• 期刊: 化学と生物 41
• 作者: Satoshi Yoshioka;et al.;由里本博也ら
• 通讯作者: 由里本博也ら

Yurimoto, H: "Alcohol dehydrogenases that catalyze methyl formate synthesis participate in formaldehyde detoxification in the methylotrophic yeast Candia boidinii"Yeast. 21(印刷中). (2004)

Yurimoto, H：“催化甲酸甲酯合成的醇脱氢酶参与甲基营养酵母 Candia boidinii 的甲醛解毒”21（出版中）。