Active site determination of novel plant versatile peroxidase extracted from Citrus sinensis and bioconversion of β-naphthol

Abstract

A ligninolytic peroxidase called versatile peroxidase, VP, (EC 1.11.1.16) is an iron-containing metalloenzyme. The most distinctive feature of this enzyme is its composite molecular framework, which combines lignin peroxidase's capacity to oxidize compounds with high-redox potential with manganese peroxidase's capacity to oxidize Mn²⁺ to Mn³⁺. In this study, we have extracted amino acid sequences from the Citrus sinensis source and subjected them to various computation tools to visualize the insight secondary and 3D structure, physicochemical properties, and validation of the structure which have not been studied so far to further investigate the catalytic efficiency and effectiveness of VP. The binding energies of HEME and HEME C (HEC) ligands with produced PDB (6rqf.1. A) have been also assessed, analyzed, and confirmed utilizing AutoDock. Binding energies were calculated using the AutoDock and validated by MD simulation using SCHRODINGER DESMOND. Most stable confirmation was achieved through a protein-ligand interaction study. Bio-technological use of VP in the biotransformation of β-naphthol has also been studied. The findings in the current study will have a substantial impact on proteomics, biochemistry, biotechnology, and possible uses of versatile peroxidase in the bio-remediation of different toxic organic compounds.

Supplementary information: The online version contains supplementary material available at 10.1007/s13205-023-03758-x.

Keywords: 3D structure; In silico; Metalloenzyme; Physicochemical; Protein–protein interaction; Versatile peroxidase.

Scheme 1

Variable peroxidase’s mechanism of substrate oxidation. Hydroperoxides oxidize the resting enzyme to a two-electron deficient compound I, which is then reduced by one electron to compound II and the natural enzyme

Fig. 1

Amino acid composition with different percentages

Fig. 2

Percentage basis composition of the different helices by using the SOMPA tool

Fig. 3

Peptide structure of the amino acid sequence by using the Pep-Draw tool

Fig. 4

(a, b) Three-dimensional structure of predicted amino acid sequence with 215 residues by using (a) SWISS-MODEL, (b) ligand-binding site prediction by using I-TASSER

Fig. 5

(a, b) Ramachandran plot of templet 6rqf.1.A by SWISS-MODEL Expasy

Fig. 6

(a, b, c) Predicted terms within the gene ontology hierarchy for molecular function, biological function, and cellular function. Confidently predicted terms are color-coded by Cscore^GO

Fig. 7

(a, b) Energy map of template 6rqf.1 drawn by using MMV software was green, sky blue, yellow, and red space representing sterically favorable, hydrogen acceptor favorable, hydrogen donor, favorable, and electrostatic, respectively

Fig. 8

RMSD of Cα atoms of protein and protein-fit ligands, and root-mean-square fluctuations (RMSF) of the protein residues of vp-HEC complex (a, b) and vp-HEME complex (c, d), respectively

Fig. 9

Normalized protein–ligand interaction bar chart and ligand–protein docking after simulation of 100 ns where (a, b) protein–HEC ligand and (c, d) protein–HEME ligand

Fig. 10

(a) Radius of gyration of HEC complex (in black color) and HEM complex (in red color). (b) Solvent-accessible surface area (SASA) of HEC-protein complex (in black color) and HEM protein complex structure complex (in red color)

Fig. 11

(a) Changes spectra of UV at 0-, 30-, and 60-min time intervals at room temperature (b) HPLC analysis of the degradation of β-naphthol of 5 mM (black line) and reaction mixture after 1 h (red line) (where, 1 ml reaction mixture contained 5 mM substrate, 50 mM buffer, 0.5 mM H₂O₂, and 1.1 mg of VP enzyme)