ERA-IB5-The exploitation of Xanthophyllomyces dendrorhous as a sustainable platform for the PROduction of high-value CARotenoids

类胡萝卜素色素在商业上用作化妆品和保健品中的生物活性成分、各种饲料中的添加剂、香料、香精和着色剂以及重要的工业原料和精细化学品。它们的工业相关性也意味着它们是具有高价值的化合物，每年的全球市场价值在10亿美元左右。迄今为止，化学合成一直是首选的生产方法。这种生产仍然很昂贵，会产生反应污染物、非天然立体异构体，并且通常与使用不可再生能源的化学炼油厂有关。类胡萝卜素有许多植物来源。然而，它们通常是次生代谢物，通常是在植物生命周期的特定发育阶段由未充分利用的、生长缓慢的和不适合现代农业实践的稀有物种在专门的细胞类型、器官或组织中产生的。植物细胞培养极易受到污染，与微生物源相比，细胞的大小限制了生物质的生产。藻类资源需要强烈的光，在下游加工过程中需要大量的能量输入，而且分子资源不够先进，无法提供有竞争力的产量。途径工程和菌株改良的现代分子方法的出现，以及发酵技术和下游加工的同步进步意味着微生物生产是植物和化学生产的有竞争力的可持续替代品。这种方法也不会与土地资源争夺粮食生产，也不会对我们日益减少的化石燃料来源产生影响。因此，微生物平台的发展为有价值化学品的可再生生物资源的创造提供了一个合乎逻辑的解决方案，具有更好的经济和环境凭证。在拟议的项目中，红酵母叶黄菌（原法菲亚红酵母）将被开发为高价值类胡萝卜素（玉米黄质和植物烯）的微生物可再生来源。在需求的推动下，这两种类胡萝卜素的市场正在迅速增长。据估计，到2018年玉米黄质的全球市场将达到3.34亿美元（BCC Research报告FOD025C）。这种对玉米黄质的兴趣与其作为治疗和预防老年性黄斑变性（AMD）的措施有关。在西方社会，慢性眼疾AMD占所有失明病例的54%。植物烯是一种无色胡萝卜素，在迅速发展的化妆品工业中被用作生物活性成分（Von Oppen-Bezalel等人，2007,SöFW-Journal 07）。它的有益活动包括防止紫外线和氧化损伤，从而导致过早老化和其他现象。作为类胡萝卜素的生产系统，黄叶菌具有天然高产香叶基焦磷酸（GGPP）衍生物萜类化合物的优势。例如，大多数细菌和酵母（如酵母菌）产生高水平的法尼酯二磷酸酯（FPP），它们不合成长链GGPP衍生的萜类物质，如类胡萝卜素。因此，合成和积累这些化合物的内在代谢和调节机制已经在这个有机体中进化。技术进步意味着黄叶菌现在有了一个测序和注释的基因组，建立了代谢组学平台，并且很容易用一套组合载体进行转化，这些组合载体能够以稳定的方式将多个基因整合到基因组中，并且可以利用农用工业废物作为碳源。积累不同的类异戊二烯前体的突变体是可用的，从实验室到工业规模的发酵程序已经建立。因此，该项目将利用及时的发展来创建一个基于微生物的平台，能够提供高价值类胡萝卜素的可再生资源。

Carotenoids pigments are used commercially as bioactives in cosmetics and health supplements, additives in a variety of feedstuffs, as fragrances, flavours and colorants as well as important industrial raw materials and fine chemicals. Their industrial relevance also means they are compounds of high-value with global markets in the range of $ 1 billion per annum. To date chemical synthesis has been the production method of choice. Such production remains expensive, creates reaction contaminants, non-native stereo isomers and is typically associated with chemical refineries using non-renewable energy sources. Numerous plant sources exist for carotenoids. However they are typically secondary metabolites often being produced in specialised cell types, organs or tissues at a specific developmental stage of the plant's life cycle by underutilised, slow growing and rare species that are not amenable to modern agricultural practices. Plant cell culture is highly prone to contamination and the size of the cells restricts biomass production in comparison to microbial sources. Algal sources require intense light, significant energy input during down-stream processing and the molecular resources are not adequately advanced to deliver competitive yields.The onset of modern molecular approaches to pathway engineering and strain improvement, with the concurrent advances in fermentation technologies and down-stream processing means microbial production is a competitive sustainable alternative to plant and chemical based production. This approach also does not compete with land resources for food production or impact on our dwindling sources of fossil fuel. Therefore the development of microbial platforms provides a logical solution to the creation of renewable biosources of valuable chemicals with improved economic and environmental credentials. In the proposed project the red yeast Xanthophyllomyces dendrorhous (formerly Phaffia rhodozyma) will be developed as a microbial based renewable source of high-value carotenoids namely zeaxanthin and phytoene. Driven by demand the markets for these two carotenoids are rapidly growing. It is estimated that the global market for zeaxanthin will reach $334 million by 2018 (BCC Research report FOD025C). This interest in zeaxanthin is associated with its use as a treatment and preventative measure against Age Related Macular Degeneration (AMD). The chronic eye disorder AMD accounts for 54% of all cases of blindness in Western societies. Phytoene is a colourless carotene used as a bioactive ingredient in the rapidly expanding cosmetic industry (Von Oppen-Bezalel et al. 2007, SöFW-Journal 07). Its beneficial activities include protection against UV and oxidative damage, which lead to pre-mature ageing and other phenomena. As a production system for carotenoids Xanthophyllomyces has the advantage of being a naturally high producer of geranylgeranyl pyrophosphate (GGPP) derived terpenoids. For example, most bacteria and yeasts (such Saccharomyces) produce high levels of farnesyl diphospate (FPP), they do not synthesis longer chain GGPP derived terpenoids such as carotenoids. Thus intrinsic metabolism and regulatory mechanisms to synthesise and accumulate such compounds have evolved in this organism. Technological advances mean that Xanthophyllomyces now has a sequenced and annotated genome, established metabolomic platforms and is readily amenable to transformation with a suite of combinatory vectors capable of integrating multiple genes into the genome in a stable manner and can utilise agro-industry waste as carbon sources. Mutants accumulating different isoprenoid precursors are available and fermentation procedures from the laboratory to industrial scale established. Thus the project will exploit timely developments to create a microbial based platform capable of delivering renewable sources of high-value carotenoids.