Isolation,identification And Degradation Mechanism Of Bisphenol A Degrading Bacteria From Deep Ocean Hydrothermal Sediments

Bisphenol A(BPA),one of the most typical endocrine disrupting chemicals(EDCs),is widely used as synthetic monomer of epoxy resin and polycarbonate in production and life.So far,BPA has been detected in most of environmental media,and cause endocrine disruption,neurotoxicity,immunotoxicity and other adverse effects on the organisms.Microbial degradation of BPA is an economic and environmental-friendly way to remove BPA,which has attracted extensive attention from experts and scholars.The hydrothermal region formed by deep ocean volcanic eruption is characterized by high pressure,high temperature,darkness and low nutrition,etc.So the microorganisms in it have unique physiological mechanism.The BPA degrading bacteria isolated from the deep ocean hydrothermal region can be used in fast and efficient degradation of BPA pollution with high salt and temperature tolerance.It provides data support for the evaluation of the potential application of deep ocean microbial resources to the environment,and expands the resource pool of BPA degrading bacteria.The main results are as follows:In this study,a strain of HQ-51-Ba with BPA degradation ability was isolated from the sediments of Atlantic deep ocean hydrothermal region.The strain was identified as Bacillus altitudinis HQ-51-Ba by 16S r RNA and genome sequencing.The single factor optimization method was used to optimize the degradation conditions of BPA.It was found that the strain HQ-51-Ba had strong adaptability to p H(6-8)and temperature(30-40^oC),and the strain had the maximum bacterial proliferation at 40^oC.Under the optimal condition,the inoculation volume of 3 m L,the glucose concentration of 0.36 g·L^-1,the p H value of6.0,and the culture temperature of 30^oC,10 mg·L^-1 BPA was completely degraded in 11 h.And UPLC-HRMS was used to analyze the intermediate products produced during the degradation of BPA.Five intermediate products of BPA were identified as 4-(2-hydroxypropyl)phenol(A),1,2-bis(4-hydroxyphenyl)-2-propanol(possibly)(C),4-hydroxyacetophenone(HAP)(E),p-hydroxybenzoic acid(HBA)(G),2-methylbutyric acid(I),and the degradation pathway of BPA by HQ-51-Ba was proposed according to the intermediate products.Genome sequencing of HQ-51-Ba was performed and submitted to Gen Bank database(Accession number:CP0407047).The sizes of strain HQ-51-Ba circular genome is 3.70Mb with GC content 41.36%and two plasmids(Plasmid A,7.206 Kb;Plasmid B,7.089Kb).After prediction,there were 3930 coding sequences(CDS),21 r RNA genes and 70t RNA genes,3131 genes were annotated into 168 KEGG pathways.Compared with previous studies,two obtained genes(cot A and bisd B)may be involved in the BPA degradation process of HQ-51-Ba.And three intermediate products were detected and identified as 4-(2-hydroxypropyl)phenol(A),1,2-bis(4-hydroxyphenyl)-2-propanol(possibly)(C),2-methylbutyric acid(I),respectively.The obtained supernatant and cell lysate degraded BPA of 10 mg·L^-1,respectively.The results that degradation efficiency of cell lysate is 54.16%in 24 h with supernatant no degradation showed that the enzymes degrading BPA were located in intracellular.Therefore,the plasmids of the two potential degradation enzymes(Thioredoxin-dependent Thiol peroxidase(TDT)and Peptidoglycan editing factor Pge F(PEF-PGEF))of BPA were selected and synthesized through genetic engineering.After transformation,induction,expression and purification,the pure enzymes were obtained,and BPA was degraded by the obtained pure enzymes.The results showed that TDT and PEF-PGEF enzyme alone could not degrade BPA,but can promote degradation of BPA by strain HQ-51-Ba.Compared with control group,the degradation efficiencies at 22 h are increased by 12.5%and 6.9%,respectively.