Recent Theoretical Insights into the Oxidative Degradation of Biopolymers and Plastics by Metalloenzymes

Abstract

Molecular modeling techniques have become indispensable in many fields of molecular sciences in which the details related to mechanisms and reactivity need to be studied at an atomistic level. This review article provides a collection of computational modeling works on a topic of enormous interest and urgent relevance: the properties of metalloenzymes involved in the degradation and valorization of natural biopolymers and synthetic plastics on the basis of both circular biofuel production and bioremediation strategies. In particular, we will focus on lytic polysaccharide monooxygenase, laccases, and various heme peroxidases involved in the processing of polysaccharides, lignins, rubbers, and some synthetic polymers. Special attention will be dedicated to the interaction between these enzymes and their substrate studied at different levels of theory, starting from classical molecular docking and molecular dynamics techniques up to techniques based on quantum chemistry.

Keywords: biofuels; biopolymers; bioremediation; laccases; lytic polysaccharide monooxygenase; molecular modeling; oxidative metalloenzymes; peroxidases; plastic; rubber oxygenases.

Figure 7

(a) Active site of a fungal laccase bound to the di-lignol model guaiacyl 4-O-5 dimer (in blue) as proposed in [168] considering 1KYA [147] Trametes versicolor laccase. The ligand is placed around 7.6 Å from the T1 Cu. Around the ligand are Phe and Ile hydrophobic interaction, the H-bond between Asp227 with one phenolic OH group. Blue arrows depict the intramolecular electron transfer from the substrate to T1 Cu and successively from T1 Cu to T2/T3 centers throughout the very conserved HCH motif. T1-T2 distance is 12–13 Å and T2-T3 3.5–4.0 Å. The substrate oxidation mechanism depends on the nature of the ligand portion under T1 attack. For phenolic portions, a H atom transfer (HAT) through Asp/Glu side chain assisted by the His residues of the T1 Cu coordination has been proposed [157,158]. For non-phenolic portions, a direct electron transfer (ET) to T1 Cu was proposed. The O₂ channel is oriented toward one of the T3 Cu centers [121]. (b) Scheme of T1 Cu site of PM1 laccase and 7D5 mutant as described in [169] with the N,N-dimethyl-p-phenylenediamine ligand (in pink). The substrate forms H-bond interactions with the His455 side chain that belongs to the T1 Cu coordination and with Asp205 side chain. The V162A mutation is evidenced and belongs to the loop that defines the binding pocket. Enlarging it, the substrate approach to T1 Cu is favored (see the blue sketch of the substrate), and thus the catalytic efficiency of the mutated enzyme is increased. The distance between the nearest carbon atom of the Val/Asp side chain is taken from PDB 5ANH (PM1) and 6H5Y (7D5) [170]. (c) T1 coordination as a function of redox potential, spanning from fungal HRPL to fungal, bacterial, and plant LMPL. This last is referred to as the laccase from Rhus vernificera. Bond distances (in Å) are obtained from the corresponding PDB.

Figure 12

(a) Structure of RoxA (PDB 4B2N) and detail on heme1 coordination. (b) Closed (left) and open (right) conformations of Lcp (PDB 5O1L and 5O1M, respectively). IMD refers to the axial imidazole ligand of heme, that is found in the 3D structure of the open state. In both states, the globin core, containing the heme active site, is colored in green-cyan, while the other three- and six-helices domains are colored in light green and purple, respectively. (c) General mechanism of dioxygenation by RoxA, RoxB, and Lcp. (d) Energetically favored pathway as calculated in [283].

Figure 1

Oxidative metalloenzymes and their polymeric substrates discussed in this review.

Figure 2

The chemical structure of the three mono-lignols. The radical–radical coupling between mono-lignols results in a variety of possible C-C and C-O ether linkages creating a complex network. The particular lignin composition in terms of mono-lignols, type of linkages, and condensed structures, such as dibenzodioxocin (5-5/β-O-4/α-O-4) and spirodienone (β-1/α-O-α), can vary from species to species and can change during the course of the plant’s growth. Baucher, Boerjan, and Ralph published comprehensive review papers in the field of lignin structure [27,30,34,35].

Figure 3

(a) Three-dimensional structure of a fungal AA9 (cellulose-active) and a bacterial AA10 (chitin-active) LPMO (PDB: 4EIS and 4ALC, respectively). The β-sandwich fold is in grey, binding-surface loops are highlighted in different colors, and copper is represented as a light blue sphere. (b) Structural detail on the active site of LPMOs depicted in panel (a) (AA9 in light blue and AA10 in yellow). The dioxygen molecule is only found in PDB 4EIS and water molecules are omitted. (c) General mechanism of LPMOs for oxidation at 1 or/and 4 positions in the presence of O₂ or H₂O₂ as a co-substrate. (d) Calculated pathways for the formation of the Cu-oxyl species according to distal or proximal protonation of the Cu-OOH⁻ intermediate.

Figure 4

(a) Three-dimensional structure of three selected LPMOs in complex with oligosaccharide substrates, namely cellopentose (left, PDB 5NLS), xylopentaose (middle, PDB 5NLO), and glucomannose (right, PDB 5NKW). (b) Interaction energy analysis (kcal/mol) for PcAA9_D (left) and HiAA9_B (right). Calculated values have been mapped on the PDB 4B5Q and 5NNS, respectively. For HiAA9_B, only one of the two pinpointed orientations is shown, and energy values have been reported as the average over the different replicas in which the selected orientation has been obtained. Horizontal orange lines indicate the orientation of the polysaccharide chains laying on the enzyme’s flat surface.

Figure 5

(a) Mechanism of the two ET from CDH to LPMO. First, the electron is transferred from DH to CYT and then from CYT to Cu(II). The structures of the closed and open conformation of CDH correspond to the PDB 4QI6 and 4QI7, respectively. (b) Proposed ET pathway in LsAA9 (PDB: 5ACG). (c) Optimized QM-portions (taken from ref [107]) of the CYT-LPMO system in the presence of oxygen before and after the ET (left) and molecular orbital occupations for the ET process in 3/2 and 1/2 state. In the 1/2 state, the electron must go directly to the O₂ molecule and the final Fe(III)Cu(I) system is found in an open-shell singlet state, which is quite unstable if compared to the triplet one.

Figure 6

(a) The crystal structure of CotA laccase found in Bacillus subtilis complexed with sinapic acid (PDB: 4Q8B). The T1 Cu site is characterized by Met502 in apical position with SMet-Cu distance of 3.26 Å, by Cys492 with S_Cys-Cu equal 2.20 Å and by His419 and His497 with N_His-Cu of 2.03 Å and 2.05 Å, respectively. T1 and T2 are connected by the His491-Cys492-His493 bridge and T1-T2 distance is 12.75 Å on average. T2-T3 Cu-Cu distances are 4.13 Å and 3.69 Å. (b) The binding site surface around the sinapic acid.

Figure 8

Laccase-mediated catalytic cycles for substrate oxidation. On the left: the direct substrate oxidation. On the right: the same substrate oxidation in the presence of a mediator.

Figure 9

(a) The crystal structure of MnP found in Phanerodontia chrysosporium (PDB: 1YYD), VP found in Pleurotus eryngii (PDB: 2BOQ), and LiP found in Phanerodontia chrysosporium (PDB: 1B82). For each enzyme, a 2D representation of the catalytically active sites is reported. (b) Scheme of the catalytic cycle reactions of class-II heme peroxidases (S represents a generic substrate).

Figure 10

Representation of the conserved residues in the heme pocket of all ligninolytic peroxidases. MnP is represented in red (PDB: 1YYD), VP is represented in blue (PDB: 2BOQ), and LiP is represented in green (PDB: 1B82).

Figure 11

(a) Representation of the conserved residues surrounding the catalytically active tryptophan in LiPs (PDB: 1B82 from P. chrysosporium). (b) Residues involved in the LRET path in LiP (right, PDB: 1B82 from P. chrysosporium) and VP (left, PDB: 2BOQ from P. eryngii).