Structural and functional studies of natural product biosynthetic enzymes

点击翻译按钮获取中文摘要

Natural products are produced by an abundance of organisms spanning all kingdoms of life. These small molecules and their derivatives are widely used in everyday living and also constitute a major source of human medicine. In recent years, sequencing of more and more microbial genomes and the genomics-guided approaches have uncovered numerous novel natural products, strongly indicating the beginning of the second golden age for the discovery of new bioactive natural products. Nature has evolved a dazzling array of enzymes to synthesize these natural products. The chemically complex structure and fascinating function of many natural products is often the result of interesting and unusual enzymatic catalysis. I propose a program on structural and functional studies of the novel and important enzymes in natural product biosynthesis which will provide mechanistic insight into the underlying logic of natural product biosynthesis. This will have potential to significantly advance our understanding of various biosynthetic pathways and offer new redesign perspectives for the generation of unnatural natural products by rational or combinatorial approaches. Our research will involve X-ray crystallographic studies of enzymes and their complexes with substrates, products and their analogues combined with functional studies using site-directed mutagenesis and enzymatic assays. We will apply acquired knowledge toward engineering new enzymes with altered specificities. In the period covered by this application, I will focus on novel enzymes in bacterial alkaloid biosynthetic pathways. Indolosesquiterpenes (ISTs), primarily found in plants, are indole-containing alkaloid natural products. Only very recently were ISTs found in prokaryotes. These molecules featuring rare pentacylic ring systems display interesting biological activities. Our specific objective is to understand how XiaI and XiaK, two novel flavin-dependent oxygenases in the xiamycin/oxiamycin biosynthetic pathway, catalyze the unusual reactions for bacterial ISTs biosynthesis. This study will facilitate protein engineering to alter the substrate profile of these enzymes, paving the way for generating novel alkaloids with improved or new biological activities.