Study On The PAHs Degradative Characteristics And The Metabolic Pathway Of Sphingomonas Sp. H

Polycyclic aromatic hydrocarbons(PAHs) are one class of organic pollutants, widely distributed in the environment.They are carcinogenic,teratogenic,and mutagenic to living organisms by food chain.It is an efficient way to recover PAHs contamination by microbial remediation.It is a hotspot field to construct Genetic Engineering Microorganisms(GEMs) capable of bioremediating contaminated environment,which denpends on the elucidation of genes and corresponding enzymes involved in PAHs degradation and therefore the clarification of genetic regulation and metabolism pathways.In this research,strain H was used to study the physiological and biochemical features,the PAHs degradative characteristics and optimize the degradation of PAHs, identifying the functional genes and enzymes involved in the biodegradation,and elucidating PAHs biodegradative pathway.The main results were as follows:1.Strain H can utilize a wide range of PAHs made of 2-4 aromatic rings as the sole carbon source and degrade these PAH compounds.Use uniform design experimentation to optimize the degradation conditions,the degradation velocity of phenanthrene(498.467 mg L^-1) was largest after 47.099 hours,the density of cells is 0.5(OD₆₀₀).2.Strain H was used to study the enzymes involved in PAHs degradation using genomic methods.The gene encoding a key enzyme was cloned and overexpressed in Escherichia coli BL21(DE3).The enzyme is catechol-2,3-dioxygenase(C23O),catalyzes the extradiol ring-cleavage of catechol.The result indicated the main catabolic pathway of strain H degrades PAHs.n c and proteomic,tion of metabolism pathways.orresponding enzymes involved in PAHs degradation and therefore the clarificat3.Two-dimensional(2D) gel electrophoresis of phenanthrene-induced proteins from cultures of strain H was used to detect proteins that increased after phenanthrene exposure.Comparison of proteins from phenanthrene-induced and uninduced cultures on 2D gels indicated that at least twelve major proteins were expressed.Proteins newly induced by phenanthrene were then analyzed from the gels by liquid chromatography/tandem mass spectrometry (LC-MS/MS).Most of them shared a high degree of similarity with the existing database and were responsible for phenanthrene or naphthalene degradation, such as hydroxylating dioxygenase,catechol dioxygenase,salicylaldehyde dehydrogenase,2-hydroxymuconic semialdehyde dehydrogenase.A complete phenanthrene degradation pathway for Sphingomonas sp.H based on this results was proposed.