2019BBSRC-NSF/BIO. SynBioSphinx: building designer lipid membranes for adaptive resilience to environmental challenges.

2019BBSRC-NSF/BIO。

• 批准号: BB/T016841/1
• 负责人: Dominic Campopiano
• 金额: $ 49.08万
• 依托单位: University of Edinburgh
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2020
• 资助国家: 英国
• 起止时间: 2020 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FT016841%2F1
• 关键词: 2019BBSRC; NSF; BIO.; SynBioSphinx; building

Animal and bacterial cells have membranes. These are protective, water-resistant shells that are composed of molecules with a water-loving (hydrophilic) head group and a long, water-hating (hydrophobic) tail. This large family of molecules are called lipids and include fats and cholesterol. One particular sub-family of lipids are sphingolipids (SLs) and ceramides which have long fatty tails. SLs sometimes have sugars attached and are known glycosphingolipids, GSLs. The SLs not only allow membranes to resist water and let nutrients in and waste out, they have also been found to stimulate the human immune system. SL levels fluctuate but are also tightly controlled. Large changes in cellular SL levels are a sign that something has gone wrong and are strongly linked with diseases such as Alzheimer's, asthma, cancer and nerve-wasting.An exciting area of research is the discovery that humans are hosts for many different types of bacteria that also make SLs, ceramides and GSLs. Collectively these bugs are known as the microbiota/microbiome and they live in our gut, on our skin and in our mouths. They are "good" bacteria - beneficial to our health. My USA collaborator recently discovered that bacteria (Caulobacter) growing in fresh water also make SLs and we are only now discovering why bacteria have such SLs. In our project we want to take advantage of SLs and use them to make membrane vesicles (like tiny soap bubbles) in a test-tube starting from basic starting materials. These vesicles are currently made chemically but a goal is to mimic nature and design cell-like, SL-containing vesicles ourselves. It is hoped that these man-made vesicles will have uses in new healthcare technologies e.g. drug delivery and detector molecules. To make the SLs we need to work in a multi-step pathway using simple building blocks. The production steps are catalysed (sped up) by molecular machines called enzymes. Research has focused on the enzymes involved in human and plant SL biosynthesis but very little is known about SL biosynthesis in bacteria. We will use these bugs as a source of the enzymes that will make SLs. If they make enough of them they will naturally come together to form synthetic vesicles. Unlike the human enzymes which need membranes to be active, the bacterial enzymes are active in water - this makes everything a lot easier, quicker and more efficient and we will make vesicles in a more controlled way. We will begin with the enzyme SPT that uses two main building blocks - an amino acid, L-serine and a long chain fatty acid, to make the first SL. We will then add one enzyme at a time to the test tube and monitor the SL formation using a technique called mass spectrometry which measures the exact weight of the molecule. As we progress the enzyme and chemistry work, my collaborators will also put the SL-producing bacteria under attack from two outside agents - an antibiotic and a bacteriophage (like a virus). The SLs in the membrane can protect them or make them more sensitive to these threats so we will use this powerful screening technique to identify the complete bacterial SL and GSL biosynthetic pathway. Then we will combine both parts of the project to pull all the enzymes together in a test tube.One scientific goal is to be able to build up designer natural and non-natural molecules in self-sufficient metabolic networks using a concept known as synthetic biology. This involves engineering concepts to design, build and test collections of biologically- and chemically-catalysed reactions. We measure the output (e.g. SLs/vesicles), learn from that process, then go around the cycle repeatedly until we find the most efficient route. It is hoped that we can use these methods to design and control life-like systems from the bottom up. The results of SynBioSphinx will be of use to academic and industrial scientists from many disciplines who are building new molecules in new ways.

动物和细菌细胞具有膜。这些是保护性的防水壳，由带有热爱水的（亲水性）头部组的分子组成，并且是长长的水，疏水（疏水）尾巴。这个大型分子家族称为脂质，包括脂肪和胆固醇。脂质的一个特定亚家族是鞘脂（SLS）和具有长脂肪尾巴的神经酰胺。 SLS有时会附着糖，并且是已知的糖磷脂，GSL。 SLS不仅允许膜抵抗水，并使营养成分并浪费出来，还发现它们刺激了人类的免疫系统。 SL水平波动，但也受到严格控制。细胞SL水平的很大变化表明出现了问题，并且与阿尔茨海默氏症，哮喘，癌症和神经浪费等疾病有着密切的联系。令人兴奋的研究领域是，人类是许多不同类型的细菌的宿主，也使SLS，Ceramides和GSLS也是如此。这些虫子共同被称为微生物群/微生物组，它们生活在我们的肠道，皮肤和嘴里。它们是“好”细菌 - 对我们的健康有益。我的美国合作者最近发现，在淡水中生长的细菌（花椰菜）也使SLS成为SLS，我们现在才发现细菌为什么具有这样的SLS。在我们的项目中，我们希望利用SLS，并使用它们从基本起始材料开始的试管中制作膜囊泡（例如小肥皂气泡）。这些囊泡目前是化学制成的，但目标是模仿本身，含有SL的囊泡。希望这些人造的囊泡能够用于新的医疗保健技术，例如药物输送和检测器分子。为了使SLS使用简单的构建块在多步途径中工作。生产步骤是由称为酶的分子机催化（加速）。研究的重点是与人类和植物SL生物合成有关的酶，但对细菌中的SL生物合成知之甚少。我们将使用这些错误作为将制造SLS的酶的来源。如果他们做到足够多，他们自然会聚在一起形成合成囊泡。与需要膜活跃的人类酶不同，细菌酶在水中活跃 - 这使得一切都变得更加容易，更快，更高效，我们将以更具控制的方式制作囊泡。我们将从使用两个主构建块的酶SPT开始 - 氨基酸，L丝氨酸和长链脂肪酸，以制造第一个SL。然后，我们将一次添加一种酶来进行试管，并使用一种称为质谱的技术来监测SL形成，该技术测量分子的精确重量。随着酶和化学工作的进行，我的合作者还将使产生SL的细菌受到两个外部药物的攻击 - 一种抗生素和一种噬菌体（如病毒）。膜中的SLS可以保护它们或使它们对这些威胁更敏感，因此我们将使用这种强大的筛选技术来识别完整的细菌SL和GSL生物合成途径。然后，我们将结合项目的两个部分，将所有酶放在试管中。一个科学目标是能够使用一种称为合成生物学的概念在自给自足的代谢网络中建立自然和非天然分子。这涉及工程概念以设计，建立和测试生物学和化学催化反应的集合。我们测量输出（例如SLS/囊泡），从该过程中学习，然后重复绕周围绕着我们找到最有效的路线。希望我们可以使用这些方法从自下而上设计和控制类似Life的系统。 Synbiosphinx的结果将用于许多以新方式建立新分子的学科的学术和工业科学家。

项目成果

Abstract 1349: Convergent evolution of bacterial ceramide synthesis

摘要 1349：细菌神经酰胺合成的趋同进化

• DOI: 10.1016/j.jbc.2023.103790
• 发表时间: 2023
• 期刊: Journal of Biological Chemistry
• 影响因子: 4.8
• 作者: Klein E