Roy J. Carver Department of Biochemistry, Biophysics and Molecular Biology, Iowa State Universitygrid.34421.30, Ames, Iowa, USA.
Department of Civil and Environmental Engineering, University of Michigan, Ann Arbor, Michigan, USA.
Appl Environ Microbiol. 2022 Jan 25;88(2):e0184121. doi: 10.1128/AEM.01841-21. Epub 2021 Nov 3.
Methanobactins (MBs) are ribosomally synthesized and posttranslationally modified peptides (RiPPs) produced by methanotrophs for copper uptake. The posttranslational modification that defines MBs is the formation of two heterocyclic groups with associated thioamines from X-Cys dipeptide sequences. Both heterocyclic groups in the MB from Methylosinus trichosporium OB3b (MB-OB3b) are oxazolone groups. The precursor gene for MB-OB3b is , which is part of a gene cluster that contains both annotated and unannotated genes. One of those unannotated genes, , is found in all MB operons and, in conjunction with , is reported to be involved in the formation of both heterocyclic groups in all MBs. To determine the function of , a deletion mutation was constructed in OB3b, and the MB produced from the Δ mutant was purified and structurally characterized by UV-visible absorption spectroscopy, mass spectrometry, and solution nuclear magnetic resonance (NMR) spectroscopy. MB-OB3b from the Δ mutant was missing the C-terminal Met and was also found to contain a Pro and a Cys in place of the pyrrolidinyl-oxazolone-thioamide group. These results demonstrate MbnC is required for the formation of the C-terminal pyrrolidinyl-oxazolone-thioamide group from the Pro-Cys dipeptide, but not for the formation of the N-terminal 3-methylbutanol-oxazolone-thioamide group from the N-terminal dipeptide Leu-Cys. A number of environmental and medical applications have been proposed for MBs, including bioremediation of toxic metals and nanoparticle formation, as well as the treatment of copper- and iron-related diseases. However, before MBs can be modified and optimized for any specific application, the biosynthetic pathway for MB production must be defined. The discovery that is involved in the formation of the C-terminal oxazolone group with associated thioamide but not for the formation of the N-terminal oxazolone group with associated thioamide in OB3b suggests the enzymes responsible for posttranslational modification(s) of the two oxazolone groups are not identical.
甲烷菌素 (MBs) 是由产甲烷菌产生的用于铜摄取的核糖体合成和翻译后修饰的肽 (RiPPs)。定义 MBs 的翻译后修饰是从 X-Cys 二肽序列形成两个具有相关硫胺的杂环基团。来自 Methylosinus trichosporium OB3b 的 MB（MB-OB3b）中的两个杂环基团都是噁唑酮基团。MB-OB3b 的前体基因是 ，它是包含注释和未注释基因的基因簇的一部分。这些未注释基因之一 ，与 一起，存在于所有 MB 操纵子中，并据报道参与所有 MB 中两个杂环基团的形成。为了确定 的功能，在 OB3b 中构建了缺失突变，并通过紫外-可见吸收光谱、质谱和溶液核磁共振 (NMR) 光谱对从 Δ 突变体中产生的 MB 进行了纯化和结构表征。从 Δ 突变体中产生的 MB-OB3b 缺少 C 末端的 Met，并且还发现含有 Pro 和 Cys 代替吡咯啉基噁唑酮-硫酰胺基团。这些结果表明，MbnC 是从 Pro-Cys 二肽形成 C 末端吡咯啉基噁唑酮-硫酰胺基团所必需的，但不是从 N 末端二肽 Leu-Cys 形成 N 末端 3-甲基丁醇-噁唑酮-硫酰胺基团所必需的。MB 已经提出了许多环境和医学应用，包括有毒金属的生物修复和纳米颗粒的形成，以及铜和铁相关疾病的治疗。然而，在 MB 可以针对任何特定应用进行修饰和优化之前，必须定义 MB 生产的生物合成途径。发现 参与与相关硫胺的 C 末端噁唑酮基团的形成，但不参与与相关硫胺的 N 末端噁唑酮基团的形成，这表明负责两个噁唑酮基团翻译后修饰的酶并不相同。
