Analysis, Predictive Modelling and In Vitro Validation of Gene Expression During 2-aminoethylphosphonate Metabolism in Sinorhizobium meliloti 1021.

苜蓿中华根瘤菌 1021 2-氨基乙基膦酸代谢过程中基因表达的分析、预测模型和体外验证。

• 批准号: EP/E057357/1
• 负责人: Nigel Ternan
• 金额: $ 6.92万
• 依托单位: University of Ulster
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2007
• 资助国家: 英国
• 起止时间: 2007 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=EP%2FE057357%2F1
• 关键词: Analysis; Predictive; Modelling; Vitro; Validation

This research project is a collaborative effort between a microbiologist and a control systems engineer who have a shared interest in examining the ways in which mathematical models can be used to predict gene expression in bacterial systems. This proposal will enable the applicants to develop expertise at the interface of their disciplines leading to new insight in the mechanisms of complex biological systems. This is expected to initiate a long-term collaborative research program beyond this project, focused on developments in the treatment and manipulation of biological systems for industrial and medical applications. For this investigation, we are using a relatively simple system, namely the metabolic route for biodegradation of an analogue of the amino acid beta-alanine, called 2-aminoethylphosphonate. This compound has a covalent carbon to phosphorus (C-P) bond that makes it very resistant to enzymatic attack. However, in the soil bacterium Sinorhizobium meliloti 1021 there is a novel pathway that allows this microorganism to use 2AEP as a source of carbon, nitrogen and phosphorus for growth. The pathway is unusual in that it generates an antiviral compound, phosphonoacetate, as an intermediate in metabolism: this is a new phenomenon, which has not been observed in bacteria until now and we wish to know more about the way in which the genes of the pathway are expressed in response to different environmental conditions. We wish to find out whether or not the five genes involved in the metabolism of 2AEP are differentially expressed in bacterial cultures that are actively growing on 2AEP. In order to do this, we will use a technique called reverse transcriptase PCR (RT-PCR) which will allow us to measure, with reference to standards of known amounts of DNA, the levels of gene expression in several samples of cells removed from cultures during growth on 2AEP. The data we will produce during this work will be of the highest quality and will therefore serve as a benchmark study in terms of both reliability and rigour of data, and should be of significant interest to systems biology researchers.A principal aim of his work is to develop modelling techniques which enable the true biochemical pathway of cellular processes to be uncovered. A major aspect of this project is therefore devoted to investigating methods for bringing together knowledge and analysis of the biochemistry with mathematical analysis. This will enable the development of a comprehensive system model, which we will then use to make predictions of the dynamics of gene expression in response to different 2AEP substrate concentrations and nutrient limitations. Validation of the feasibility of the proposed model is extremely important and following rigorous simulation tests, further targeted gene expression analysis will be carried out on bacterial cultures fed with small amounts of 2AEP in order to test the in silico predictions. The biological implications of the model will be studied in terms of understanding of the responses of soil microbes to nutrient influx; determining the cellular control mechanisms; and robustness of the system to environmental noise. The research will be of interest to scientists interested in metabolism of organophosphonate compounds, which are widely used in medicine, agriculture and industry. The techniques developed will also be of wider interest to others in the field of modelling biochemical pathways dynamics. The techniques could be of use in predicting gene expression behaviour in similar systems, including perhaps those related to pathogenesis of certain bacteria and this could provide a starting point for the development of new antimicrobials. The investigators will benefit significantly from the work, with the reciprocal scientific interchange broadening their understanding of the ways by which systems biology experiments may be fed into and used to design predictive models of metabolism.

该研究项目是微生物学家和控制系统工程师之间的合作成果，他们对研究数学模型用于预测细菌系统中基因表达的方法有共同的兴趣。该提案将使申请人能够在其学科的界面上发展专业知识，从而在复杂生物系统的机制中获得新的见解。预计这将启动一项长期的合作研究计划，重点关注工业和医疗应用中生物系统的治疗和操作的发展。在这项研究中，我们使用了一个相对简单的系统，即生物降解氨基酸β -丙氨酸类似物的代谢途径，称为2-氨基乙基膦酸盐。这种化合物有一个共价碳磷键（C-P），使其对酶的攻击具有很强的抵抗力。然而，在土壤细菌Sinorhizobium meliloti 1021中，有一种新的途径允许这种微生物利用2AEP作为碳、氮和磷的来源进行生长。该途径的不寻常之处在于，它在代谢过程中产生一种抗病毒化合物——磷酸乙酸盐，这是一种新的现象，直到现在还没有在细菌中观察到，我们希望更多地了解该途径的基因是如何在不同的环境条件下表达的。我们希望了解参与2AEP代谢的5个基因在2AEP上活跃生长的细菌培养物中是否存在差异表达。为了做到这一点，我们将使用一种称为逆转录酶PCR （RT-PCR）的技术，该技术将使我们能够根据已知DNA量的标准来测量在2AEP上生长期间从培养物中取出的几个细胞样本中的基因表达水平。我们将在这项工作中产生的数据将是最高质量的，因此将作为数据可靠性和严谨性的基准研究，并且应该引起系统生物学研究人员的极大兴趣。他工作的一个主要目的是开发建模技术，使细胞过程的真正生化途径得以揭示。因此，该项目的一个主要方面是致力于研究将生物化学的知识和分析与数学分析结合起来的方法。这将有助于开发一个全面的系统模型，然后我们将使用该模型来预测基因表达对不同2AEP底物浓度和营养限制的响应动态。验证所提出模型的可行性非常重要，在严格的模拟测试之后，将对少量2AEP喂养的细菌培养物进行进一步的靶向基因表达分析，以测试计算机预测。该模型的生物学意义将在理解土壤微生物对养分流入的反应方面进行研究；确定细胞控制机制；以及系统对环境噪声的鲁棒性。该研究将引起对有机膦化合物代谢感兴趣的科学家的兴趣，这些化合物广泛应用于医学，农业和工业。开发的技术也将对生化途径动力学建模领域的其他人产生更广泛的兴趣。这些技术可以用于预测类似系统中的基因表达行为，包括可能与某些细菌的发病机制有关的系统，这可以为开发新的抗菌剂提供一个起点。研究人员将从这项工作中受益匪浅，互惠的科学交流拓宽了他们对系统生物学实验可能被输入并用于设计代谢预测模型的方式的理解。

项目成果

A SYSTEMS BIOLOGY APPROACH TO UNDERSTANDING THE PATHWAY BY WHICH A BIOGENIC ORGANOPHOSPHONATE IS PRODUCED DURING METABOLISM OF 2AEP IN S. meliloti.

一种系统生物学方法，用于了解苜蓿草中 2AEP 代谢过程中产生生物有机磷酸盐的途径。

• 发表时间: 2008
• 期刊: Book of Abstracts
• 作者: N G Ternan