(Below N is a link to NCBI taxonomic web page and E link to ESTHER at designed phylum.) > unclassified entries: NE > unclassified sequences: NE > metagenomes: NE > metagenome: NE
LegendThis sequence has been compared to family alignement (MSA) red => minority aminoacid blue => majority aminoacid color intensity => conservation rate title => sequence position(MSA position)aminoacid rate Catalytic site Catalytic site in the MSA MGGGGSGGGNNGGGGGCEADCGYERGPDPSVSLLEASTGPFSVRTSNVSS SVRGFGGGTIHYPTNTTGTMAAIVVIPGFVSPESSIAWWGPKLASHGFVV MTIGTNSGFDQPASRASQLNNALDYLIEQNGSSRSPINGMIDTDRLGVMG WSMGGGGTLRVATEGRVSAAIPLAPWDSSSSQFRSIDTPTLIFACENDST APVRSHADPFYDAIPDSTAKAFVELDGGGHTCANGSSGFGGSYNDVLSRL GVSWMKLHLDKDQRYNQFVCGPNHESDRSISEYRGTCPYLE
Polyethylene terephthalate (PET) is one of the most abundantly produced synthetic polyesters. The vast number of waste plastics including PET has challenged the waste management sector while also posing a serious threat to the environment due to improper littering. Recently, enzymatic PET degradation has been shown to be a viable option for a circular plastic economy, which can mitigate the plastic pollution. While protein engineering studies on specific PET degradation enzymes such as leaf-branch compost cutinase (LCC), Thermobifida sp. cutinases and Ideonella sakaiensis PETase (IsPETase) have been extensively published, other homologous PET degrading enzymes have received less attention. Ple629 is a polyester hydrolase identified from marine microbial consortium having activity on PET and the bioplastic polybutylene adipate terephthalate (PBAT). In order to explore its catalytic mechanism and improve its potential for PET hydrolysis, we solved its crystal structure in complex with a PET monomer analogue, and validated its structural and mechanistic similarity to known PET hydrolases. By structural comparisons, we identified some hot spot positions described in previous research on protein engineering of PET hydrolases. We substitute these amino acid residues in Ple629, and obtained variants with improved activity and thermo-stability. The most promising variant D226A/S279A exhibited a more than 5.5-fold improved activity on PET nanoparticles than the wild-type enzyme, suggesting its potential applicability in the biotechnological plastic recycling.
Polybutylene adipate terephthalate (PBAT) is a biodegradable alternative to polyethylene and can be broadly used in various applications. These polymers can be degraded by hydrolases of terrestrial and aquatic origin. In a previous study, we identified tandem PETase-like hydrolases (Ples) from the marine microbial consortium I1 that were highly expressed when a PBAT blend was supplied as the only carbon source. In this study, the tandem Ples, Ple628 and Ple629, were recombinantly expressed and characterized. Both enzymes are mesophilic and active on a wide range of oligomers. The activities of the Ples differed greatly when model substrates, PBAT-modified polymers or PET nanoparticles were supplied. Ple629 was always more active than Ple628. Crystal structures of Ple628 and Ple629 revealed a structural similarity to other PETases and can be classified as member of the PETases IIa subclass, alpha/beta hydrolase superfamily. Our results show that the predicted functions of Ple628 and Ple629 agree with the bioinformatic predictions, and these enzymes play a significant role in the plastic degradation by the consortium.
The degradation of synthetic polymers by marine microorganisms is not as well understood as the degradation of plastics in soil and compost. Here, we use metagenomics, metatranscriptomics and metaproteomics to study the biodegradation of an aromatic-aliphatic copolyester blend by a marine microbial enrichment culture. The culture can use the plastic film as the sole carbon source, reaching maximum conversion to CO(2) and biomass in around 15 days. The consortium degrades the polymer synergistically, with different degradation steps being performed by different community members. We identify six putative PETase-like enzymes and four putative MHETase-like enzymes, with the potential to degrade aliphatic-aromatic polymers and their degradation products, respectively. Our results show that, although there are multiple genes and organisms with the potential to perform each degradation step, only a few are active during biodegradation.
Marinobacter Hydrolase (PETase-like) Cold-Adapted PET Hydrolase Ple629
