Construction Of Genome-scale Metabolic Model For Halomonas Sp.TD01

Genome-scale metabolic models(GSMMs)have been universally acknowledged as the indispensable tool in biological researches,which can comprehensively investigate and analyze the metabolism of microbial cells from a systematic perspective,and build a bridge from identifying the basic components of cells to understanding the complex growth phenotype.As a gram-negative and moderate halophile,Halomonas sp.TD01 was endowed with dominant characteristics,such as the strong adaptability to the environment,the rapid growth with simple nutritional demands and so forth.Moreover,it possesses the capacity of accumulating polyhydroxyalkanoates(PHA)at high content while with low-cost renders it more preferences.This study aims to provide a powerful and reliable GSMMs analysis tool for acquiring comprehensive insights on the metabolic behavior of Halomonas sp.TD01 and enormously assisting in designing metabolic engineering strategies for developing high value-added chemicals.Above all,we utilized a semi-automated construction tool to construct the first genome-scale metabolic model of Halomonas sp.TD01 based on genome annotation,database information and related literatures.By reviewing several literatures,the basic criteria and specific method flow of the unified and standardized treatment for model format,the judgment and revision of reaction direction and reversibility as well as the identification,analysis and filling of Gaps was summarized in detail.A set of efficient and convenient inspection procedure was designed for the four common ATP generation errors,the quality control of the model was then carried out from the perspective of energy generation,and the optimal ATP production rate was 202.5mmol/g DW/h.Then,the biomass reaction was modified.Due to the lack of reference data of cell composition,the divided compositions with their mass fraction of each building block were mostly obtained and adapted from C.salexigens i FP764,and the i AF1260of E.coli was also acknowledged as an auxiliary reference.Meanwhile,taking the specificity of TD01 accumulating compatible solutes into account,the biomass equation composed of 81 precursors was generated in the end.Through verifying and analyzing the integrity and correctness of all the biomass precursor synthesis pathways,the optimal synthesis specific-rate of biomass was finally obtained to be0.995 h^-1.Ultimately,the substrate profiling of TD01 was obtained and analyzed by Biolog Phenotype Micro Array(PM)experiment,and the results demonstrated that TD01exhibited a wide-ranging substrate spectrum and showed signal response with different intensities to 129 of the 190 test carbon source substrates.Based on these growth phenotype data,the i XZ1051 model was checked,analyzed and modified,ultimately obtaining great performance improvement[(from the original 48.95%(93/190)to 88.95%(169/190)]and proving its accuracy from a side view.In the final model verification stage,the predicted outcomes gained from quantitative simulations under distinct conditions demonstrated high consistency with that determined in previous experiments,indicating i XZ1051 possesses excellent capabilities on elucidating the basic metabolism of Halomonas sp.TD01.A series of tedious process with iteratively manual refinement,verification,modification and optimization finally led to i XZ1051 encompassing 1911 reactions,1698 metabolites and 1051 genes.Therefore,we expected that the i XZ051 model present here can be leverages as a high-quality platform to explore the biotechnological potential of Halomonas sp.TD01 and further facilitate future studies.