Molecular Evolution and Adaptation of Microbial Polyketide Biosynthetic Sequences

洛佩兹聚酮(PK)化合物及其生物合成酶代表了微生物中不同的表型，但这种多样性的潜在原因尚不完全清楚。本研究的主要假设是生物合成聚酮合成酶(PKS)序列多样性和程序化是由正向多样化选择驱动的，并解释了自然界观察到的大部分聚酮代谢物多样性。将采用比较生物信息学的方法来实现以下目标：a)通过与脂肪酸合成酶(FAS)的比较或通过使用Firebird等生物信息学软件来推断已发表的PKS基因座的二级和高级酶结构，例如编码红霉素、雷帕霉素和利福霉素的那些；b)比较“类似”模块的序列和预测的酶结构(在单一PK途径内)，并将这些与相应的PK代谢物或PK中间体相关联，并推断基因复制的可能性；C)比较(不同细菌和不同途径之间)同源的PKS酶模块序列和结构，以获得与PK代谢物最终产物的相关性，并调查水平基因转移的可能性和频率；d)如果PKS结构在结构上看起来同源，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，而如果结构不同源，则使用预测的结构构象的数据调查活性中心如何保持功能以及为什么它们是保守的；e)确定是否积极地选择了对结构确定和整体酶功能至关重要的密码子和残基(以保存类似/同源结构或功能)；F)从以前从HBOI收集中鉴定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)的生物合成相结合。公众影响大自然在许多层面上表现出其多样性，其中之一是天然产品的生物化学多样性。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品用于实际目的。然而，在遗传序列数据和它们的表型(基因的物理表达)之间存在着很大的知识差距，本研究旨在填补这一差距。尽管如此，分子序列信息仍可用于跟踪进化历史，推断分子结构，并获得某些编码聚酮天然产物的基因的当代生态背景。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并不理想。这项研究可以对聚酮化合物的供应问题产生直接影响，因为研究的目标是参与聚酮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.