Fig. 2
From: Functional tailoring of a PET hydrolytic enzyme expressed in Pichia pastoris
The effects of N-glycosylation on P. pastoris-expressed CtPL-DM. a SDS-PAGE analysis of recombinant proteins expressed in E. coli and secreted by P. pastoris with or without EndoH treatment. Lane M: protein marker. The theoretical migration distance of CtPL-DM on SDS-PAGE is indicated by an arrow. b PET hydrolytic activity of CtPL-DM and each variant expressed by E. coli and P. pastoris that were treated with or without EndoH. The hydrolytic products (MHET and TPA) released from GfPET film by each enzyme as well as blank control without enzyme were measured at 18 h. A triplicate assay was conducted and the average values ± standard deviation are presented. UD, undetectable; 3A, N181A/N220A/N261A. c Crystal structure of CtPL-DM (PDB ID, 8IAN), N-glycosylation sites are indicated as yellow sticks and transparency spheres with glycan colored by elements. The catalytic triad residues (black dashed circle) are shown as sticks. d The zoom-in views of four putative N-glycosylation sites