Table 1
Fitness Score and Interacting amino acids of lead compounds from Blighia sapida and control ligand.
Compound Name | PubChem ID | Fitness Score | H-Bond Residues | Interacting active site hydrophobic amino acids |
Quercetin | 5280343 | 1.865 | MET 140, ASP 138 | ILE 61, MET 140, LEU 139, LEU 137, ALA 82, LEU 189, ILE 115, TYR 66, VAL 69 |
Kaempferol | 5280863 | 1.498 | LYS 84, ASN 187, ASP 138 | TYR 66, VAL 69, ALA 82, MET 140, LEU 139, LEU 137, ILE 115, LEU 189 |
(+)-catechin | 9064 | 1.705 | LYS 84, ASP 138, ASN 187 | VAL 69, TYR 66, ALA 82, MET 140, LEU 139, LEU 137, ILE 115, LEU 189, |
Actaealactone | 11537736 | 1.809 | ASP 138, GLU 102. OTHER INTERACTIONS: TYR 66(PI-PI STACKING), LYS 84(SALT BRIDGE) | MET 140, LEU 139, LEU 137, ILE 115, ALA 82, PHE 201, TYR 66, LEU 189, VAL 69 |
Co-crystallized ligand | 118959080 | 2.428 | MET 140, ASP 138. OTHER INTERACTIONS: TYR 66(PI-PI STACKING) | LEU 189, MET 140, LEU 139, |
Alloathyriol | 44575387 | 1.52 | ASP 138, ASP 200. OTHER INTERACTIONS: LYS 84(SALT BRIDGE) | VAL 69, TYR 66, LEU 106, PHE 201, ALA 82, ILE 115, LEU 189, LEU 137, LEU 139, MET 140, ILE 61 |
Molecular Docking, Drug-like Properties, And Interaction Profiling Of Erk5-ligand Complexes
For molecular docking, computational approaches are frequently employed to predict the ligand-receptor complex structure; this is commonly accomplished by sampling ligand conformations in the protein's active site and ranking the conformations.
Table 2
Drug Likeness properties of lead compounds and control ligand
Entry Name | mol MW | Hbond Acceptors | Hbond Donors | ALogP | Polar Surface Area | Rule of Five |
Quercetin | 302.24 | 7 | 5 | 2.531 | 131.36 | 0 |
Kaempferol | 286.24 | 6 | 4 | 2.7984 | 111.13 | 0 |
(+)-Catechin | 290.272 | 6 | 5 | 1.9202 | 110.38 | 0 |
Actaealactone | 358.304 | 7 | 4 | 0.9343 | 147.35 | 0 |
Co-crystallized Ligand | 388.195 | 4 | 1 | 2.7951 | 71.95 | 0 |
Alloathyriol | 274.229 | 5 | 2 | 1.7616 | 102.96 | 0 |
The interaction of the ligands inside the binding pocket of Extracellular signal-regulated kinase 5 (ERK5), MM-GBSA(Table 1), and their pharmacokinetic profile are all part of this computational analysis (Table 6). ERK5 is a protein kinase with a transcriptional transactivation domain and a nuclear localization signal(Cook et al., 2020). Because ERK5 inhibition has therapeutic potential in cancer and inflammation, ERK5 kinase inhibitors have been developed( Carmell et al., 2021).
From Quercetin to Alloathyriol, bioactive compounds from Blighia sapida demonstrated a favorable binding affinity and optimally saturated the active site of ERK5, with binding energies of -9.257kcal/mol and 7.847kcal/mol, respectively. A stronger binding is associated with lower binding energy. The lead compounds in Table 1 bind securely inside the active region of ERK5 while creating primary amino acid interactions with the hydrophobic amino acid residues MET 140, LEU 189, LEU 139, TYR 66, LEU 137, ALA 82, ILE 115, and VAL 69, according to the docking technique (Table 1; Fig. 5 ). These amino acid residues are critical for anticipating the ERK5 binding site and catalytic mechanism. Through H-bond formation with the hydroxyl group, the docked molecules interact with MET 140, ASP 138, ASP 200, GLU 102, ASN 187, and LYS 84 in the ERK5 binding pocket.
Inter and intramolecular interactions such as hydrogen bonding, pi-pi stacking, pi-cation, and salt bridge occur as a result of the ligand-ERK5 complexes. The best bioactive molecule of Blighia sapida was quercetin, which had maximum binding energy of 9.257kcal/mol. Hydrogen bonds are formed between it and the hydrophobic and negatively charged amino acids MET 140 and ASP 138. With binding energy of 8.788kcal/mol, kaempferol is thought to interact mostly with negative-charged, positive-charged, and polar amino acids: LYS 84, ASN 187, and ASP 138. (+)-Catechin is found to completely occupy the ERK5 binding site with a binding energy of 8.384kcal/mol while interacting with negative, positive, and polar amino acids: LYS 84, ASP 138, and ASN 187 via hydrogen bonds. While establishing a hydrogen bonding connection with negatively charged amino acid, ASP 138, actaealactone, and alloathyriol had binding energies of 7.974 and 7.847kcal/mol, respectively(Fig. 5). They both had additional interactions, including a salt bridge connection with LYS 84, a positively charged amino acid(Fig. 6). In addition, Actaealactone and TYR 66 developed a PI-PI stacking interaction. The top four bioactive compounds of Blighia sapida have higher binding energy than the pyrole inhibitor (4-(2-bromanyl-6-fluoranyl-phenyl)carbonyl-N-pyridin-3-yl-1H-pyrrole-2-carboxamide) that was utilized as a positive control for the ligands. The binding energy of the pyrolle inhibitor was 7.864kcal/mol, indicating that Blighia sapida bioactive molecules have a strong potential to bind ERK5 for the treatment of various cancers, particularly breast cancer.
The MM-GBSA module, which is integrated with the Schrodinger suite's prime program, was used to compute the ∆Gbind for ERK5-lead ligand complexes. Following the docking analysis, the ∆Gbind was used to calculate the binding energy for the screened compounds using advanced mechanics. MM-GBSA technique is a reliable post docking method for estimating the binding position of docked complexes, according to several researchers. Quercetin, Kaempferol, (+)-Catechin, Actaealactone, and Alloathyriol had binding energies of 36.10, 53.04, 44.67, 29.59, and 24.16 kcal/mol, respectively, according to the MM-GBSA output (Fig. 4).
DENSITY FUNCTIONAL THEORY ANALYSIS
Thermochemical analysis
Thermodynamic characteristics are important factors in determining the spontaneity and chemical stability of a chemical reaction. Gibbs free energy is a thermodynamic parameter used to characterize the interaction between ligands and receptors. It shows the likelihood of biomolecular processes taking place. A positive free energy value suggests that binding will not occur until additional external energy is added, whereas a negative free energy value indicates that binding will occur spontaneously. The degree of the negative free energy determines the extent of the ligand's interaction with the receptor. Enthalpy is also a measure of a thermodynamic system's total energy. When the ligand attaches to the receptor, the binding enthalpy indicates the energy change in the system. The computed Gibb's free energy for the investigated compounds is shown in Table 4. Because all of the molecules have a negative free energy, they can attach to the target receptor without requiring any external energy. The top hit chemical, Quercetin, as well as Acetalactone, had the highest free energy (-1103.987 and 1295.839Hartree, respectively), implying that Acetalactone will interact more than the other compounds investigated. The dipole moment reveals the polarity of a compound as well as the distribution of electrons inside it(Fleming, 1977). It improves the receptor protein's binding affinity, non-bonded interactions, and hydrogen bond formation. Acetalactone(7.00debye) also has the largest dipole moment, as demonstrated in Table 4.
Table 4
Molecular weight, electronic energy, enthalpy, Gibb’s free energy and Dipole moment values of Blighia sapida Lead compounds obtained via DFT at the B3LYP/6-31G* level.
Compounds | Molecular Weight | Electronic Energy | Enthalpy | Gibbs Free Energy | Dipole Moment |
Quercetin | 302.238 | -1104.18 | -1103.929 | -1103.987 | 0.22 |
Kaempferol | 286.239 | -1028.96 | −1028.717 | −1028.773 | 1.54 |
(+) – Catechin | 290.271 | -1031.33 | -1031.042 | -1031.101 | 3.56 |
Acetalactone | 358.302 | -1296.12 | -1295.77 | -1295.839 | 7.00 |
Alloathyriol | 274.228 | -990.86 | -990.62 | -990.679 | 2.14 |
Frontier molecular orbital (FMOs)
The most important orbitals in the molecule are the FMOs, HOMO, and LUMO. They are important in optical, electric, and UV-Vis spectral chemistry, as well as quantum chemistry. The FMOs describe how a molecule interacts with other molecules and provide information on electron transport in a molecule, as well as a molecule's chemical reactivity and stability. The HOMO energy defines the molecule's propensity to donate electrons; greater EHOMO values imply a stronger inclination for the molecule to donate electrons(44). The ELUMO determines a molecule's ability to receive an electron; a lower ELUMO number improves the likelihood of taking electrons. Therefore, higher values of EHOMO and lower values of ELUMO are responsible for the low stability and high reactivity of a molecule. The EHOMO values for the compounds tested rise in order as shown in Table 5: Quercetin > Kaempferol > (+) – Catechin > Alloathyriol > Acetalactone. The greatest value of EHOMO is found in quercetin (5.48eV), indicating that it has a higher tendency to donate an electron to the target receptor than other substances. In addition, the calculated ELUMO values are provided in Table 5. Quercetin has the lowest ELUMO value, indicating that it has a lower ability to receive electrons than the other compounds investigated. Furthermore, as shown in Table 3, both the EHOMO and ELUMO are completely dispersed across the molecule structure, implying considerable HOMO-LUMO overlapping, resulting in robust charge transfer behavior. The band gap energy between the EHOMO and ELUMO is crucial for forecasting a molecule's chemical reactivity. The chemical reactivity and stability of a molecule are reflected in the band gap energy levels. The molecule becomes tougher, more stable, and less reactive as the band gap energy increases. High reactivity and low stability are associated with a narrowing of the energy band gap. The energy band gap values are as follows: Quercetin < Kaempferol < Alloathyriol < Acetalactone < (+) – Catechin. Among the isolated chemicals, quercetin has the smallest band gap, indicating that it is more reactive toward the target receptor than the others.
Global reactivity descriptors
Table 5
Global reactivity descriptors of Blighia sapida lead compounds obtained via DFT.
Compounds | EHOMO (eV) | ELUMO (eV) | Eg (eV) | I((eV) | A(eV) | η (eV) | δ (eV− 1) | χ (eV) | µ(eV− 1) |
Quercetin | -5.48 | -1.84 | 3.64 | 5.48 | 1.84 | 1.82 | 0.549451 | 3.66 | -3.66 |
Kaempferol | -5.53 | -1.81 | 3.72 | 5.53 | 1.81 | 1.86 | 0.537634 | 3.67 | -3.67 |
(+) -Catechin | -5.63 | -0.08 | 5.71 | 5.63 | -0.08 | 2.855 | 0.350263 | 2.775 | -2.775 |
Acetalactone | -6.02 | -1.78 | 4.24 | 6.02 | 1.78 | 2.12 | 0.471698 | 3.9 | -3.9 |
Alloathyriol | -5.74 | -1.55 | 4.19 | 5.74 | 1.55 | 2.095 | 0.477327 | 3.645 | -3.645 |
(Eg), energy band gaps; (EHOMO), highest occupied molecular orbital energy; (ELUMO), lowest unoccupied molecular orbital energy; (I), ionization energy; (δ), chemical softness; A, electron affinity; η, chemical hardness; µ, chemical potential; χ, electronegativity. |
To gain a thorough understanding of the chemical stability and reactivity of bioactive chemicals toward the target receptor, global reactivity descriptors (GRD) were developed. Ionization energy, electron affinity, chemical hardness, chemical softness, chemical potential, and electronegativity are the GRDs that are calculated. The ionization energy (I) of a molecule describes its chemical reactivity and stability. It's the amount of energy it takes to remove an electron from a molecule. High ionization energy suggests chemical inertness and stability, whereas low ionization energy indicates high reactivity and chemical inertness(Chakraborty et al., 2020). The ionization energy of quercetin (5.48 eV) is the lowest, making it the most reactive chemical toward the target receptor, ERK5The energy released when an electron is added to a neutral molecule is known as electron affinity (A). A molecule with a high electron affinity is more likely than one with a low electron affinity to receive electrons easily(Geerlings & Proft, 2002). The most reactive chemicals include Kaempferol and Quercetin, which have the highest electron affinity.
Understanding the reactivity of a chemical system requires an understanding of chemical hardness and softness. Chemical hardness describes a molecule's resistance to electron cloud deformation (Mortier et al., 1985). The band gap energy of a hard molecule is enormous, whereas the band gap energy of a soft molecule is tiny. The soft molecule will polarize more quickly and easily than the hard molecule (Obot et al., 2015). Catechin has the greatest hardness value (2.86eV) in Table 5, suggesting that it is the hardest molecule. The softest chemical is quercetin, which has the lowest softness value (0.54eV). The ability of a molecule to draw electron electrons toward itself is known as electronegativity. Table 5 shows that acetalactone is the most electronegative of all the chemicals (3.9eV).
Evaluation of ADMET and druglikeness properties of lead compounds
Table 6
Pharmacokinetics profile of lead compounds of Blighia sapida
Model | Quercetin | Kaempferol | (+)-Catechin | Actaealactone | Alloathyriol |
QPlogHERG | -4.875 | -5.054 | -4.897 | -4.223 | -4.476 |
QPPCaco | 19.934 | 54.93 | 49.958 | 10.054 | 218.753 |
QPlogBB | -2.305 | -1.824 | -1.95 | -2.673 | -1.186 |
QPPMDCK | 7.185 | 21.49 | 19.395 | 3.429 | 95.701 |
QPlogKhsa | -0.354 | -0.201 | -0.427 | -0.597 | -0.257 |
% Oral Absorption | 52.196 | 64.083 | 59.884 | 42.501 | 76.2 |
QPlogKhsa: Binding to human serum albumin (− 1.5 to + 1.5) |
QPlogHERG: IC50 value for blockage of HERG K + channels (below − 5) |
QPPMDCK: Apparent Madin-Darby canine kidney cell permeability in nm/sec. |
QPlogBB: Brain/blood partition coefficient (− 3.0 to 1.2) |
QPPCaco: Apparent Caco-2 cell permeability in nm/sec (< 25 poor, > 500 great) |
The pharmacokinetics of phytochemicals is important to know before their development into new pharmaceutical drugs. Using the QikProp module of Schrodinger, pharmacokinetic properties of the lead compounds were predicted using binding to human serum albumin (QPlogkhsa), Madin-Darby canine kidney cell permeability (QPPMDCK), blood/brain partition coefficient (QPlogBB), IC50 value for the blockage of HERG K + channels (QPlogHERG) and Caco-2 cell permeability (QPPCaco). All of the lead compounds fell into the range of values for QPlogKHSA, therefore they should all be able to bind to human serum albumin. All of the lead compounds except Kaempferol have been predicted to have good values for QPlogHERG. Again, all of the lead compounds fell within the range of expected values for Brain/Blood partition coefficient. The results also showed that only Quercetin and Actaealactone have poor Caco-2 cell permeability.