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Future directions in cofactor research: Discovery and development of coenzyme F420-dependent bioprocesses

辅因子研究的未来方向：辅酶 F420 依赖性生物过程的发现和开发

基本信息

批准号：
408113938
负责人：
Professor Dr. Gerald Lackner
金额：
--
依托单位：
Abteilung Molekulare und Angewandte Mikrobiologie
依托单位国家：
德国
项目类别：
Research Grants
财政年份：
2018
资助国家：
德国
起止时间：
2017-12-31 至 2020-12-31
项目状态：
已结题

来源：
https://gepris.dfg.de/gepris/projekt/408113938?language=en
关键词：
Future directions cofactor research Discovery

项目摘要

Cofactors (coenzymes) are helper molecules that are crucial for the catalytic activity of many enzymes. While some cofactors are ubiquitous in nature, others are found in microbes with a highly-specialized metabolism. Coenzyme F420, for instance, plays a physiological role in methanogenic microbes, antibiotic-producing bacteria, as well as in the tuberculosis pathogen. F420-dependent enzymes are highly interesting for biocatalysis, since they are able to promote challenging chemical reactions. Despite these extraordinary features, F420 remains underexplored and barely used in biotechnology. One reason for this is the low availability of the cofactor as chemical synthesis is cumbersome and microbial sources of the cofactor are non-satisfactory at the moment. Therefore, our goal is to identify a missing step in the biosynthesis of F420 to produce it in the model bacterium Escherichia coli. Production titers will then be increased by rational genetic engineering as well as directed evolution. To effectively use cofactors, a regeneration system is required that resets the cofactor to its original state after reaction. Some do exist for F420, however, all of them suffer from severe limitations. Thus, we will establish an efficient regeneration system based on inexpensive substrates. These efforts will render F420 more attractive for biotechnological applications and will facilitate further work packages.In particular, we aim to identify novel F420-dependent bioprocesses with potential use for biotechnology. Our preliminary results corroborate recent bioinformatics studies that suggest that F420-dependent enzymes are more widespread than anticipated. Exploiting their ability to bind to F420, these enzymes will be enriched from various bacteria and tested for academically intriguing and biotechnologically useful activities. Additionally, we will investigate F420 in unusual ecological systems. To this end, we will address the symbiosis between bacteria and a mold fungus, the causative agent of rice seedling blight. Our preliminary results demonstrate that F420 is produced as soon as symbionts live in association with their fungal host. In collaboration, we want to elucidate the function of F420 in this model bacterial-fungal alliance that is of high importance for microbial ecology and plant pathology.

辅因子（辅酶）是对许多酶的催化活性至关重要的辅助分子。虽然一些辅因子在自然界中普遍存在，但其他辅因子存在于具有高度专业化代谢的微生物中。例如，辅酶F420在产甲烷微生物、产甲烷细菌以及结核病病原体中起着生理作用。F420依赖性酶对于生物催化非常有趣，因为它们能够促进具有挑战性的化学反应。尽管F420具有这些非凡的特性，但它仍然没有得到充分的研究，几乎没有在生物技术中使用。其中一个原因是辅因子的低可用性，因为化学合成是繁琐的，并且辅因子的微生物来源目前不令人满意。因此，我们的目标是确定F420生物合成中缺失的步骤，以在模式细菌大肠杆菌中产生它。然后，通过合理的基因工程以及定向进化来提高生产滴度。为了有效地使用辅因子，需要在反应后将辅因子重置为其原始状态的再生系统。有些确实存在F420，但是，所有这些都受到严重的限制。因此，我们将建立一个高效的再生系统的基础上廉价的基板。这些努力将使F420在生物技术应用中更具吸引力，并将促进进一步的工作包。特别是，我们的目标是确定新的F420依赖的生物过程与生物技术的潜在用途。我们的初步结果证实了最近的生物信息学研究表明，F420依赖酶比预期的更广泛。利用它们与F420结合的能力，这些酶将从各种细菌中富集，并进行学术上有趣和生物技术上有用的活动测试。此外，我们将研究F420在不寻常的生态系统。为此，我们将讨论细菌和霉菌之间的共生关系，霉菌是水稻秧苗枯萎病的病原体。我们的初步研究结果表明，F420的产生，只要共生体生活在与他们的真菌宿主。在合作中，我们希望阐明F420在这种模式细菌-真菌联盟中的功能，这对微生物生态学和植物病理学非常重要。