Evolved P450 Mutants for Streamlining Synthesis via Late-stage C-H Activation

通过后期 C-H 激活进化 P450 突变体以简化合成

• 批准号: 1809776
• 金额: --
• 依托单位: University of Oxford
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2016
• 资助国家: 英国
• 起止时间: 2016 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-1809776
• 关键词: Evolved; P450; Mutants; Streamlining; Synthesis

The ability to effect selective hydroxylation of a specific C-H bond has profound implications for chemical synthesis and drug discovery. This proposal will further develop an ongoing collaborative effort to apply evolved cytochrome P450s in this important endeavour. We have shown that a panel of >100 unique bacterial P450 mutants developed within the Wong group converts a remarkably diverse set of organic substrates into (usually) mono-hydroxylated derivatives. Neutral lipophilic, polar, and charged molecules are oxidised, e.g. lidocaine, naproxen, and steroids such as testosterone. The general scheme for these oxidations underlies the enzymes' remarkable reactivity; reactions at 100mg substrate, routinely convert at catalyst loadings of 0.1 mol% or less. The only requirement is for a C-H bond to be physically accessible to the ferryl intermediate, and un-activated C-H bonds may therefore be oxidised. In contrast, chemical reagents show a complete preference for allylic and benzylic C-H bonds, and heteroatoms. P450-mediated oxidations are easy to conduct, require no special handling, employ mild conditions with no heavy metals and are tolerant of heteroatoms (including N and S), clean, sustainable and safe.Our research within this area falls under three headings: i) Achieving 'difficult' oxidations; ii) Facilitating target synthesis; and iii) Medicinal chemistry applications. The initial object of this proposal concerns the second heading with application to the development of novel, selective herbicides. Viridiol, a furanosteroid secondary metabolite found in a handful of fungal strains, is potentially interesting as a new herbicide but there are concerns surrounding its mammalian toxicity that must be addressed. Viridiol has not yet been synthesised but the closely related antifungal kinase inhibitor viridin has been the subject of a few synthetic studies, including one total synthesis.This molecule provides an ideal test-case to show how retrosynthetic considerations can be 're-programmed' based on a dual skeleton-building/late-stage oxidation strategy. Here, we will be free to construct the furanosteroid core by short skeleton-building sequences, carrying through minimal oxy-functionality thus dispensing with protecting group regimes. With the pentacyclic furanosteroid core prepared, research will begin to explore mutant P450 oxidation at both the activated (benzylic) and un-activated sites around the periphery, including the benzenoid ring-C. All the obtained viridiol analogues will be tested for herbicidal activity and in mammalian toxicity models to provide a structure-activity-relationship profile. This project falls within the EPSRC Physical Sciences research area.

对特定的C-H键进行选择性羟基化的能力对化学合成和药物开发具有深远的意义。这项提议将进一步发展正在进行的合作努力，将进化的细胞色素P450应用于这一重要努力。我们已经证明，在Wong集团内开发的一组&gt；100独特的细菌P450突变体可以将一组非常多样化的有机底物转化为(通常)单羟基衍生物。中性的亲脂性、极性和带电分子被氧化，例如利多卡因、萘普生和类固醇，如睾酮。这些氧化反应的一般方案是基于酶的显著反应活性；底物为100 mg的反应通常在催化剂负载量为0.1mol%或更少的情况下进行转化。唯一的要求是C-H键在物理上可以被铁基中间体访问，因此未活化的C-H键可能被氧化。相反，化学试剂表现出对烯丙基和苄基C-H键以及杂原子的完全偏好。P450介导的氧化反应易于进行，不需要特殊处理，使用温和的条件，不含重金属，对杂原子(包括N和S)具有耐受性，清洁、可持续和安全。我们在这一领域的研究分为三个方面：i)实现困难的氧化；ii)促进目标合成；iii)药物化学应用。这项建议的最初目的涉及第二个标题，即应用于开发新的选择性除草剂。Viridiol是一种在少数真菌菌株中发现的呋喃类固醇次级代谢物，作为一种新的除草剂具有潜在的兴趣，但围绕其哺乳动物毒性的担忧必须得到解决。Viridiol尚未合成，但密切相关的抗真菌激酶抑制剂viridin已经成为几项合成研究的主题，其中包括一项全合成。该分子提供了一个理想的测试案例，以展示如何基于双重骨架构建/后期氧化策略对逆合成考虑进行重新编程。在这里，我们将自由地通过短的骨架构建序列来构建呋喃类固醇核心，进行最低限度的氧合官能化，从而省去了保护基团的制度。随着五环呋喃类固醇核心的制备，研究将开始探索突变体P450在外围活化(苯基)和非活化位置的氧化，包括苯环-C。所有获得的病毒二醇类似物都将在哺乳动物毒性模型中进行除草活性测试，以提供结构-活性-关系曲线。该项目属于EPSRC物理科学研究领域。