BioDiscovery Institute and Department of Biological Sciences, University of North Texas, Denton, Texas, USA.
mSphere. 2024 Sep 25;9(9):e0049624. doi: 10.1128/msphere.00496-24. Epub 2024 Aug 27.
Methanotrophic bacteria play a vital role in the biogeochemical carbon cycle due to their unique ability to use CH as a carbon and energy source. Evidence suggests that some methanotrophs, including , can also use CO as a carbon source, making these bacteria promising candidates for developing biotechnologies targeting greenhouse gas capture and mitigation. However, a deeper understanding of the dual CH and CO metabolism is needed to guide methanotroph strain improvements and realize their industrial utility. In this study, we show that expresses five carbonic anhydrase (CA) isoforms, one α-CA, one γ-CA, and three β-CAs, that play a role in its inorganic carbon metabolism and CO-dependent growth. The CA isoforms are differentially expressed, and transcription of all isoform genes is induced in response to CO limitation. CA null mutant strains exhibited markedly impaired growth compared to an isogenic wild-type control, suggesting that the CA isoforms have independent, non-redundant roles in metabolism and physiology. Overexpression of some, but not all, CA isoforms improved bacterial growth kinetics and decreased CO evolution from CH-consuming cultures. Notably, we developed an engineered methanotrophic biocatalyst overexpressing the native α-CA and β-CA with a 2.5-fold improvement in the conversion of CH to biomass. Given that product yield is a significant cost driver of methanotroph-based bioprocesses, the engineered strain developed here could improve the economics of CH biocatalysis, including the production of single-cell protein from natural gas or anaerobic digestion-derived biogas.IMPORTANCEMethanotrophs transform CH into CO and multi-carbon compounds, so they play a critical role in the global carbon cycle and are of interest for biotechnology applications. Some methanotrophs, including , can also use CO as a carbon source, but this dual one-carbon metabolism is incompletely understood. In this study, we show that carbonic anhydrases are critical for this bacterium to optimally utilize CO. We developed an engineered strain with improved CO utilization capacity that increased the overall carbon conversion to cell biomass. The improvements to methanotroph-based product yields observed here are expected to reduce costs associated with CH conversion bioprocesses.
产甲烷菌在生物地球化学碳循环中起着至关重要的作用，因为它们具有将 CH 作为碳和能源源的独特能力。有证据表明，一些产甲烷菌，包括 ，也可以将 CO 作为碳源，这使得这些细菌成为开发针对温室气体捕获和缓解的生物技术的有前途的候选者。然而，需要更深入地了解 CH 和 CO 的双重代谢，以指导产甲烷菌菌株的改进并实现其工业应用。在这项研究中，我们表明 表达五种碳酸酐酶 (CA) 同工型，一种α-CA、一种γ-CA 和三种β-CAs，它们在其无机碳代谢和 CO 依赖性生长中发挥作用。CA 同工型的表达不同，并且所有同工型基因的转录都在 CO 限制下被诱导。与同基因野生型对照相比，CA 缺失突变菌株的生长明显受损，这表明 CA 同工型在 代谢和生理学中具有独立的、非冗余的作用。一些（但不是全部）CA 同工型的过表达改善了细菌的生长动力学，并减少了消耗 CH 的培养物中 CO 的释放。值得注意的是，我们开发了一种过表达天然α-CA 和β-CA 的工程化甲烷生物催化剂，将 CH 转化为生物质的转化率提高了 2.5 倍。鉴于产物产率是甲烷生物过程的重要成本驱动因素，因此这里开发的工程菌株可以提高基于甲烷生物的生物工艺的经济性，包括从天然气或厌氧消化衍生的沼气生产单细胞蛋白。重要性 产甲烷菌将 CH 转化为 CO 和多碳化合物，因此它们在全球碳循环中起着至关重要的作用，并且对生物技术应用感兴趣。一些产甲烷菌，包括 ，也可以将 CO 作为碳源，但这种双重一碳代谢尚不完全清楚。在这项研究中，我们表明 碳酸酐酶对于该细菌最佳利用 CO 至关重要。我们开发了一种具有改进的 CO 利用能力的工程菌株，该菌株提高了整体碳转化为细胞生物质的效率。这里观察到的产甲烷菌产物产率的提高预计将降低与 CH 转化生物工艺相关的成本。
