Discovery and Genetic Code Expansion of a Polyethylene Terephthalate (PET) Hydrolase from the Human Saliva Metagenome for the Degradation and Bio‐Functionalization of PET

Abstract

We report a bioinformatic workflow and subsequent discovery of a new polyethylene terephthalate (PET) hydrolase, which we named MG8, from the human saliva metagenome. MG8 has robust PET plastic degradation activities under different temperature and salinity conditions, outperforming several naturally occurring and engineered hydrolases in degrading PET. Moreover, we genetically encoded 2,3-diaminopropionic acid (DAP) in place of the catalytic serine residue of MG8, thereby converting a PET hydrolase into a covalent binder for bio-functionalization of PET. We show that MG8(DAP), in conjunction with a split green fluorescent protein system, can be used to attach protein cargos to PET as well as other polyester plastics. The discovery of a highly active PET hydrolase from the human metagenome-currently an underexplored resource for industrial enzyme discovery-as well as the repurposing of such an enzyme into a plastic functionalization tool, should facilitate ongoing efforts to degrade and maximize reusability of PET.

Efficient depolymerization of polyethylene terephthalate (PET) and polyethylene furanoate by engineered PET hydrolase Cut190

“…Most recently, Pfaff et al (2022) constructed L136F/Q138Y mutant of PES-H1 as the best performing variant. Although the importance of the Q138 position in PET hydrolases has no doubt, type I (TfCut2, Cut190 and LCC) and type II (IsPETase, PES-H1) hydrolases (Eiamthong et al 2022) showed different replacement effects. Considering these results, we made several Cut190 variants, as listed in Table 1 and compared their PET hydrolyzing activities (Table 4).…”

“…Most recently, a novel PET hydrolase was cloned from a compost metagenome (Sonnendecker et al 2022) and its variant indicated the comparable activity to the LCC variant (Pfaff et al 2022). In addition, PET hydrolases are cloned even from human saliva metagenome (Eiamthong et al 2022). Thermostable mutants of PETase from mesophilic Ideonella sakaiensis (Yoshida et al 2016) have been documented so far.…”

Capturing Covalent Catalytic Intermediates by Enzyme Mutants: Recent Advances in Methodologies and Applications

“…The Dap-containing PETase can also carry other cargos to functionalize PET as well as other polyester plastics. [42]…”

Mechanoenzymatic reactions for the hydrolysis of PET

“…Although PET is particularly resistant to biodegradation, 3,4 a number of naturally occurring PET hydrolases have been reported. [5][6][7][8] Amongst these are cutinases from Fusarium solani, [9][10][11][12] a leaf-branch metagenome (LCC) 6,[13][14][15] and Humicola insolens (HiC) 11,16,17 as well as a hydrolase from Thermobida fusca (TfH). 9,10,18,19 In 2016, the rst known bacterium capable of using PET as its sole carbon source, Ideonella sakaiensis (Is), was discovered.…”