Exploring, evolving and exploiting coupled racemase/acylase biotransformation systems.

探索、发展和利用耦合消旋酶/酰基酶生物转化系统。

基本信息

批准号：
BB/K006487/1
负责人：
Dominic Campopiano
金额：
$ 22.17万
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：
2013
资助国家：
英国
起止时间：
2013 至无数据
项目状态：
已结题

来源：
https://gtr.ukri.org/projects?ref=BB%2FK006487%2F1
关键词：
Exploring evolving exploiting coupled racemase

项目摘要

Modern medicine has used drugs to cure disease, alleviate chronic pain and increase life spans. Drug companies must make drugs under very strict regulations - these have to be 100% pure. One complication is that Nature has evolved to make some chemicals look the same, weigh the same, are made of the exact same atoms but they are different in the fact that they are mirror images of each other - like a pair of hands - these are called "enantiomers". It turns out that Nature tends to work with only one enantiomer and it is often observed that the opposite one is toxic, this is the case with drugs, we desire only one enantiomer. Important types of natural molecules where handed-ness make a big impact are amino acids. These are the building blocks of the proteins inside every cell. Proteins are made up hundreds and thousands of amino acids, polymerised together head-to-tail like beads on a string. These chains fold up into specific 3 dimensional shapes that can carry out many essential functions in the cell. Enzymes are also proteins and they are the workhorses of the cell - enzymes are tiny catalysts that speed up the conversion of molecule A to molecule B. Without an enzyme these conversions would take years but an enzyme catalyst can accelerate the speed of a reaction over 10 billion times. Enzymes allow us to breakdown our food, provide us with energy and help us repair damaged tissue. It turns out that these enzymes can also be put to work to make the very molecules that drug companies want. Enzymes are very specific and only work with one particular hand/mirror image of a molecule. Pharmaceutical companies endeavour to make large amounts of drugs the cheapest, purest and least wasteful way they can. Drugs are complicated molecules, many are made from amino acid building blocks (only one mirror image) in a multi-step process. Because the process uses only one of the mirror images of the starting material, the other mirror image is not used and in thus 50% is wasted.Our project aims to tackle this fundamental problem. We aim to make key amino acid building blocks of only one hand or another and use up 100% of the starting material. We will use enzymes to carry out the conversion of amino acid precursors to the target amino acid. The enzymes themselves were not designed for this specific job so we have to engineer the enzymes at a molecular level. We can do this by rational design - with knowledge of the molecular structure we can make specific changes and hope that the new enzyme will have the desired characteristics - speed, efficiency and stability. We can also carry out a random approach then fish out the desired new enzyme from the mixture. The enzyme we study catalyses the interconversion of the mirror image of one amino acid precursor into the other mirror image - this is called a racemase. Once we have the ideal racemase we will pair it up with another enzyme - an acylase - this one converts the amino acid precursor into the final amino acid but is specific for only one of the mirror images. So, we will start with both starting amino acid precursors - 50% of each mirror image. The acylase will convert one half into the product until it is used up; at the same time the racemase will be doing its job converting the unused precursor into its mirror image and when this happens the acylase can convert it. In a perfect world all of the precursor will be used up (100% conversion) and there will be no precursors left. Moreover, the enzymes can be produced cheaply, re-cycled, are bio-degradable and they work in water. We have already made good progress and now require funds to optimise the whole process. We will do this at University in partnership with a company that are experts in making amino acid precursors and products for the pharmaceutical industry. As well as making valuable tools for drug production we will also gain fundamental knowledge about enzyme design that others can apply to numerous useful processes.

现代医学已经使用药物来治疗疾病、减轻慢性疼痛和延长寿命。制药公司必须在非常严格的规定下生产药品——这些药品必须是 100% 纯的。一个复杂的问题是，大自然已经进化到使一些化学物质看起来相同，重量相同，由完全相同的原子组成，但它们的不同之处在于它们是彼此的镜像 - 就像一双手 - 这些是称为“对映体”。事实证明，大自然倾向于只使用一种对映体，并且经常观察到相反的一种对映体是有毒的，药物就是这种情况，我们只需要一种对映体。氨基酸是手性产生重大影响的重要天然分子类型。这些是每个细胞内蛋白质的组成部分。蛋白质由成百上千个氨基酸组成，像串珠一样从头到尾聚合在一起。这些链折叠成特定的 3 维形状，可以在细胞中执行许多基本功能。酶也是蛋白质，它们是细胞的主力 - 酶是微小的催化剂，可以加速分子 A 向分子 B 的转化。如果没有酶，这些转化将需要数年时间，但酶催化剂可以将反应速度加快 10 倍以上十亿次。酶使我们能够分解食物，为我们提供能量并帮助我们修复受损的组织。事实证明，这些酶也可以用来制造制药公司想要的分子。酶是非常专一的，只能与分子的一个特定的手/镜像一起工作。制药公司致力于以最便宜、最纯净和最少浪费的方式生产大量药物。药物是复杂的分子，许多是由氨基酸构件（只有一个镜像）通过多步骤过程制成的。由于该过程仅使用起始材料的一个镜像，因此不会使用另一个镜像，因此浪费了 50%。我们的项目旨在解决这一基本问题。我们的目标是仅用一只手或另一只手制造关键氨基酸构建块，并使用 100% 的起始材料。我们将使用酶将氨基酸前体转化为目标氨基酸。酶本身并不是为这项特定工作而设计的，因此我们必须在分子水平上对酶进行工程设计。我们可以通过合理设计来做到这一点——借助分子结构的知识，我们可以进行特定的改变，并希望新的酶具有所需的特性——速度、效率和稳定性。我们还可以采用随机方法，然后从混合物中找出所需的新酶。我们研究的酶催化一种氨基酸前体的镜像向另一种镜像的相互转化——这称为消旋酶。一旦我们有了理想的消旋酶，我们将其与另一种酶（酰基转移酶）配对，这种酶将氨基酸前体转化为最终的氨基酸，但仅对其中一个镜像具有特异性。因此，我们将从两个起始氨基酸前体开始 - 每个镜像的 50%。酰化酶会将一半转化为产物，直至用完；同时，外消旋酶将完成其工作，将未使用的前体转化为其镜像，当发生这种情况时，酰基转移酶可以将其转化。在完美的世界中，所有前体都会被用完（100% 转化），并且不会留下任何前体。此外，这些酶可以廉价生产、回收、可生物降解，并且可以在水中发挥作用。我们已经取得了良好的进展，现在需要资金来优化整个流程。我们将在大学与一家专门为制药行业生产氨基酸前体和产品的公司合作开展这项工作。除了为药物生产制造有价值的工具外，我们还将获得有关酶设计的基础知识，其他人可以将其应用于许多有用的过程。