Fig. 6: Comprehensive structural analysis of three novel esterases and functional validation of site-directed mutant.

A The 3D structure of Phylogenetic tree of three newly mined proteins, plotted using the chiplot online website (https://www.chiplot.online/). B proteins NOD_1, OCA_1, and OCB_1 expected three-dimensional structures. The colors in the model represent the predicted local distance difference test (pLDDT) confidence scores, with very high (>90), high (80), medium (70), low (60), and very low (<50) levels depicted in red, yellow, green, light blue, and blue, respectively. C The binding pockets of protein NOD_1, OCA_1, and OCB_1 are displayed, and the yellow area is the binding pocket site predicted by PrankWeb. D Sequence identification maps of recombinant proteins NOD_1, OCA_1, and OCB_1. The height of the letters indicates the base frequency at each location, and the abscissa represents the site of the amino acid. This figure was generated using WebLogo 3 (https://weblogo.threeplusone.com/). E SDS-PAGE of recombinant bacteria E. coli BL21-pET28a- OCA_1 mutant strain. Lane 1 was marker, and lane 2 was the protein of the E. coli BL21-pET28a- OCA_1 mutant strain purified by nickel column, and the band size was 31 kd, which was consistent with the expected protein size. F The histogram showed whether the mutant strain protein of OCA_1 had a degrading effect on 12 macrolide antibiotics. Data are given as mean ± SD, n = 3. G It reflects the susceptibility of the recombinant bacterium E. coli BL21-pET28a-OCA_1-S102 mutant to 12 macrolide antibiotics. The experiment was performed in triplicate.