Molecular Evolution and Adaptation of Microbial Polyketide Biosynthetic Sequences

LopezPolyketide （PK）化合物及其生物合成酶在微生物中表现出多种表型，但这种多样性的根本原因尚不完全清楚。本研究的主要假设是生物合成聚酮合成酶（PKS）序列多样性和“编程”是由正向多样化选择驱动的，占自然界中观察到的聚酮代谢物多样性的大部分。比较生物信息学方法将实现以下目标：A)通过与脂肪酸合成酶（FAS）的比较或使用生物信息学软件（如FIREBIRD），推断已发表的PKS位点的二级和高阶酶结构，例如红霉素、雷帕霉素和利福霉素的编码；b)比较同源模块（在单一PK通路内）的序列和预测的酶结构，并将其与相应的PK代谢物或PK中间体相关联，推断基因重复的可能性；c)比较同源（不同细菌和不同途径之间）PKS酶模块序列和结构，获得与PK代谢物终产物的相关性，并研究基因水平转移的可能性和频率；d)如果PKS结构在结构上是同源的，确定初级序列中对结构确定至关重要的特定残基和密码子，而如果结构不是同源的，研究活性中心如何保持功能，以及为什么它们是保守的，使用预测结构构象的数据；e)确定对结构决定和整体酶功能至关重要的密码子和残基是否被积极选择（以保持类似/同源结构或功能）；f)从HBOI收集的海洋微生物中分离出几个新的或更长的PKS基因序列片段。通过这些分析，将建立一个基于PKS酶序列、结构和功能进化的分子适应模型，并将其与最终表型产物（PK代谢物）的生物合成结合起来。自然的多样性体现在许多层面上，其中一个是天然产物的生化多样性。在细胞、遗传和分子水平上了解这种多样性的基本基础，将有助于将聚酮类抗生素等天然产物用于实际用途。然而，基因序列数据与其表型（基因的物理表达）之间存在很大的知识差距，本研究旨在填补这一空白。然而，分子序列信息可以用于跟踪进化历史，推断分子结构和获得某些基因编码的聚酮类天然产物的当代生态背景。此外，尽管已经发现了许多具有治疗潜力的天然化合物，但这些产品的供应往往不是最佳的。这项研究可以直接影响聚酮类化合物的供应问题，因为研究的目标是参与聚酮类化合物生物合成的基因。

LopezPolyketide (PK) compounds and their biosynthetic enzymes represent diverse phenotypes in microorganisms, but the underlying causes for this diversity are not completely understood. The major hypothesis of this research is that biosynthetic polyketide synthase (PKS) sequence diversity and "programming" is driven by positive diversifying selection and accounts for the majority of polyketide metabolite diversity observed in nature. A comparative bioinformatics approach will be implemented to achieve the following objectives: a) Infer the secondary and higher order enzyme structures of published PKS loci, such as those coding for erythromycin, rapamicin, and rifamycin by comparisons with fatty acid synthases (FAS) or by using bioinformatics software such as FIREBIRD; b) Compare sequences and predicted enzyme structures of "paralogous" modules (within a single PK pathway) and correlate these with corresponding PK metabolite or PK intermediates and infer possibility of gene duplications; c) Compare orthologous (between different bacteria and different pathways) PKS enzyme module sequences and structures to obtain correlations with PK metabolite end products and also to investigate the possibility and frequency of horizontal gene transfer; d) If PKS structures appear structurally homologous, determine specific residues and codons in primary sequences which are pivotal for structural determination, while if structures are not homologous, investigate how active centers remain functional, and why they are conserved, using data on predicted structural conformations; e) Determine whether codons and residues which appear pivotal for structure determination and overall enzyme function are positively selected (to preserve analogous/homologous structures or function); f) Isolate several new or longer PKS gene sequence segments from marine microbes previously characterized from the HBOI collection. Through these analyses, a model of molecular adaptation based on the evolution of PKS enzyme sequence, structure, and function will be formulated and integrated with the biosynthesis of the ultimate phenotypic products (PK metabolites). Public ImpactNature manifests its diversity at numerous levels, with one being the biochemical diversity of natural products. Understanding the fundamental bases of this diversity, at cellular, genetic and molecular levels, will facilitate the harnessing of natural products, such as polyketide antibiotics, for practical purposes. However a large knowledge gap exists between genetic sequence data and their phenotypes, the physical expression of genes, which this research aims to fill. Still, molecular sequence information can be used for tracking the evolutionary history, inferring molecular structures and obtaining the contemporary ecological context of certain genes coding for polyketide natural products. Moreover, despite the many natural compounds with therapeutic potential that have been discovered, the supply of those products is often less than optimal. This research can have a direct impact on polyketide supply issues since the target of study are the genes involved in the biosynthesis of polyketide compounds.