Development of Biocatalysts for Fluorination of Organic Compounds

描述（由申请人提供）：1970年，5%的许可药品含有至少一个氟原子。2005年，这一数字急剧上升至20%，据估计，目前25%的管道药物现在含有至少一个氟原子。氟对药物发现和功能的重要性怎么强调都不过分。然而，化学蒸馏的工业方法代表了目前实践的一些最危险和毒性的化学方法，通常使用NaNO 2、HF和用F2直接蒸馏。相比之下，自然界以酶促反应的典型高选择性并且在温和的水性环境中进行水解。“氟化酶”生物催化剂作为化学合成的补充方法的应用提供了制备具有生物医学重要性的重要氟化化合物的更安全、更环境可接受的途径的潜力。我们提出了一种新的高通量筛选方法，以确定和改善新的微生物来源的氟化酶生物催化剂。作为最初的目标，我们将集中在氟酶，最近在链霉菌cattleya已被证明进行C-F键的形成，但目前拥有的活性太低，是工业上有用的。我们将证明这种酶的定向进化和鉴定的进一步的新的fluorinases从基因组文库作为分子生物学方法的原理证明，以提供有用的生物催化剂，可以开发用于工业发酵的改进。氟化酶还将用作将F-18同位素掺入适当生物分子中的合成工具，特别是鉴于正电子发射断层扫描（PET）作为诊断技术的快速增长以及作为通过临床试验评估所有药物的生物分布的要求。 最近发现的微生物氟酶将无机氟离子转化为有机氟。氟的生物催化结合为清洁和安全的有机氟生物合成以及将放射性标记的氟[F-18同位素]结合到药物中开辟了巨大的生物技术机会。对氟化酶适用性的广泛兴趣源于对含有氟原子的精细化学品、医药和农业化学实体的巨大商业兴趣，以及F-18同位素在正电子发射断层扫描（PET）应用中用于医学成像的日益增长的应用。针对提高这种酶的效率和效用的研究是独特的，并且随后会有许多应用。

DESCRIPTION (provided by applicant): In 1970 5% of licensed pharmaceuticals contained at least one fluorine atom. In 2005 this figure had risen dramatically to 20% and it is now estimated that 25% of current pipeline drugs now contain at least one fluorine atom. The importance of fluorine to pharmaceutical discovery and functionality cannot be overstated. However, industrial methods of chemical fluorination represent some of the most hazardous and toxic chemical methods currently practiced, generally utilizing NaNO2, HF and direct fluorination with F2. Nature, in contrast carries out fluorination with the typically high selectivity of enzymatic reactions and in a mild aqueous environment. The application of "fluorinase" biocatalysts as a complementary approach to chemical synthesis offers the potential of safer, more environmentally acceptable routes to prepare important fluorinated compounds of biomedical importance. We propose to apply a novel high-throughput screening approach to identify and improve new fluorinase biocatalysts from microbial sources. As the initial target we will focus upon a fluorinase, recently identified in Streptomyces cattleya which has been shown to carry out C-F bond formation but which currently possesses activity too low to be industrially useful. We will demonstrate the improvement of this enzyme by directed evolution and the identification of further novel fluorinases from genomic libraries as the proof-of-principle of molecular biological approaches to provide useful biocatalysts which can be developed for industrial fluorination. The fluorinase will be applied also as a synthetic tool for the incorporation of the F-18 isotope into appropriate bio- molecules, particularly in light of the rapid growth in positron emission tomography (PET) as a diagnostic technique and as a requirement for the assessment of the biodistribution of all drugs progressing through clinical trials. The recently discovered microbial fluorinase enzyme converts inorganic fluoride ion to organic fluorine. Biocatalytic incorporation of fluorine opens up a tremendous biotechnological opportunity for clean and safe organofluorine biosynthesis and also the incorporation of radio-labeled fluorine [F-18 isotope] into drugs. The widespread interest in the fluorinase's applicability arises from the huge commercial interest in the fine chemicals, medicinal and agrochemical entities that contain fluorine atoms and also in the growing application of the F-18 isotope in positron emission tomography (PET) applications for medical imaging. The research directed towards improving the efficiency and utility of this fluorination enzyme is unique and numerous applications would follow.