The Degradation Effect Of Bacillus Velezensis On Polyethylene And Its Degradation Genes Analysis

Plastic products with good chemical stability and low production cost are widely used in various industries of manufacturing life.Post-consumer polyethylene(PE)accumulates in the environment in large quantities,causing unavoidable pollution,due to its high production demand and short service life.To address the two key problems of poor bioaccessibility of PE plastics and unclear key degradation enzymes,this thesis carried out the screening and excavation of efficient PE degradation strains.A strain of Bacillus velezensis C5 with PE degradation potential was used to characterize PE biodeterioration and biodepolymerization effect by qualitative and quantitative methods.By analyzing the whole genome sequence of B.velezensis C5,key PE biodegradation genes/ enzymes were mined,and PE biodegradation pathways and degradation mechanisms were resolved,which provided an important research basis for enzyme engineering modification.The main results of this thesis are as follows.(1)In this thesis,a novel bacterial strain C5(CGMCC No.1.18715)with low-density polyethylene(LDPE)degradation potential was obtained by selective cultivation of LDPE as the sole carbon source to screen for strains with enhanced LDPE bioaccessibility using landfill soil as the source of the strain.This strain biodegraded untreated LDPE.Based on its 16 S r RNA sequence,this strain was identified as Bacillus velezensis,a useful addition to the library of PE degrading strains.(2)The study of B.velezensis C5 in PE biodeterioration showed that the rapid biofilm formation of C5 bacteria could improve the bioaccessibility of LDPE.By analyzing hydrophobicity evaluation,surface morphology and functional groups,B.velezensis C5 was able to utilize untreated LDPE as the only carbon source to grow and rapidly form biofilm.The bioaccessibility of LDPE was significantly improved by B.velezensis C5,which decreased its contact angle from 100 ° to 54 ° within 3 days.A large amount of proteins and polysaccharides were secreted by B.velezensis C5,which adhered to the surface of LDPE extensively,increased the effective contact between enzymes and LDPE,and promoted the direct interaction between enzymes and LDPE.LDPE continuously biodeteriorated and its thickness was reduced by 26% in90 days with the help of B.velezensis C5 biofilm.(3)The study of B.velezensis C5 in LDPE biodepolymerization showed that B.velezensis C5 degraded low chain polymers to promote LDPE biodepolymerization.By analyzing the weight loss,molecular weight change,depolymerization intermediates and short-chain alkane degradation,LDPE was further biodepolymerized after B.velezensis C5 biodeterioration.Under the action of B.velezensis C5,the mass of LDPE appeared to gain weight with biofilm adhesion and gradually decreased after bio-depolymerization occurred,achieving a mass loss of about 8% in 90 days.The weight-average molecular weight of LDPE polymer increased by 29.8% and the polymer dispersity index increased by 9.4%,indicating that B.velezensis C5 depolymerized the terminal or branching low molar mass chains of LDPE and released C24-C29 short chain n-alkane intermediates.By the analysis of short-chain alkane degradation,B.velezensis C5 had the ability to degrade short-chain alkanes rapidly,with 86.74% degradation of dodecane and 67.99% degradation of cetane in 3 days.(4)The analysis of key degradation genes/ enzymes of B.velezensis C5 in LDPE biodegradation showed that B.velezensis C5 achieved multi-enzyme co-catalytic degradation of LDPE polymer with the assistance of biofilm.By sequencing the whole genome of B.velezensis C5,there were 3755 coding genes.During the biodegradation of LDPE,B.velezensis C5 expressed a cascade of degradation enzymes related to colonization,oxidation and hydrolysis to complete the catalytic degradation.Alkane compounds are transported through the sulfur metabolic pathway of B.velezensis C5,followed by assimilation and mineralization through the fatty acid degradation pathway.The genome mining results suggest a possible biofilm-assisted degradation mechanism that may involve key enzymes,such as laccase,cytochrome P450 and propionyl-Co A carboxylase,which could constitute a multi-enzyme system to co-catalyze the degradation of LDPE waste.This study expands the strain library of efficient LDPE degrading bacterium,improves LDPE bioaccessibility,and provides a useful reference for resolving key enzymes and biodegradation pathways for LDPE degradation.It lays a good foundation for the subsequent enzyme engineering modification,which helps to realize the recycling of carbon resources of waste plastics and promote the realization of carbon peaking and carbon neutrality goals.