Structural Elucidation and Biosynthesis

在上一个资助期，我们的酶学和合成化学研究补充了遗传和生物化学的努力，以了解天然产物膦酸盐的生物合成逻辑。该计划项目组已经取得了重大进展，开发了有效的方法来检测新的膦酸酯生物合成基因簇，并详细了解了导致商业上重要的膦酸酯（如膦丝菌素和磷霉素）的生物合成途径。随着遗传和生物化学研究变得更加有效，发现了一个新的瓶颈，其涉及低效的纯化策略，因此减慢了新发现的基因簇产物的结构解析。该分项目中的部分拟议研究将侧重于开发有效的净化方法， 这些新化合物的结构解析。此外，在过去的资助期内，基因组测序工作确定了相同的天然产物在不同的生物体中通过非常不同的生物合成途径产生的情况。我们将研究这些趋同进化的例子。最后，许多天然产物膦酸酯以短肽形式产生，以促进靶生物体中肽渗透酶的摄取。而对于一些化合物，小的非核糖体肽合成酶（NRPS）簇是 许多含膦酸酯的肽以不同的、通常未鉴定的方式组装，这是形成肽键的原因。我们将试图提供更多的洞察力，这一非常重要的方面方面，未来可能的工程努力。

In the previous grant period, our enzymology and synthetic chemistry studies complemented the genetic and biochemical efforts towards understanding the biosynthetic logic of natural product phosphonates. The program project team has made significant progress, developing effective methods to detect new phosphonate biosynthetic gene clusters and understanding in great detail the biosynthetic pathways leading towards commercially important phosphonates such as phosphinothricin and fostomycin. As the genetic and biochemical studies became more efficient, a new bottleneck was identified that involves inefficient purification strategies and hence slow-down of structure elucidation of the products of newly discovered gene clusters. Part of the proposed studies in this subproject will focus on the development of efficient methods for purification and structural elucidation of those new compounds. In addition, the genome sequencing efforts of the past grant period identified cases in which the same natural product is made in different organisms by very different biosynthetic routes. We will investigate these examples of convergent evolution. Finally, many natural product phosphonates are produced as short peptides to promote uptake by peptide permeases in the target organisms. Whereas for some compounds small non-ribosomal peptide synthetase (NRPS) clusters are responsible for peptide bond formation, many phosphonate containing peptides are assembled in different, often unidentified ways. We will attempt to provide more insight into this very important aspect with respect to possible future engineering efforts.