key: cord-0776182-us9rwsic
authors: Liu, Ju-Zhao; Lv, Hong-Chang; Fu, Yu-Jie; Cui, Qi
title: Amomum tsao-ko essential oil, a Novel Anti-COVID-19 Omircon Spike Protein Natural Products: A Computational Study
date: 2022-04-18
journal: Arab J Chem
DOI: 10.1016/j.arabjc.2022.103916
sha: 5a107807d1cddf343bbdd225e397f0aa9ab87323
doc_id: 776182
cord_uid: us9rwsic

Since the outbreak of COVID-19, this virus has been constantly mutating. The latest mutant Omicron has been identified as VOC by WHO. The main reason for its concern is the mutation of 46 amino acids in spike protein, which has brought the global epidemic prevention into another difficulty. Herbal aromatic plant Amomum tsao-ko was excavated from formula 1 and 2 for the treatment of COVID-19 in China, and its active components were extracted and identified. Molecular dynamics simulation and Fpocket were applied to find the key sites on RBD(Omicron), and molecular docking was also used to reveal the interaction between A. tsao-ko essential oil (AEO) and RBD(Omicron). The AEO components were analyzed and identified by GC/Q-TOF MS. There were 20 kinds of AEO with a relative area percentage of more than 1%, and they were related to the three active centres of RBD(Omicron). In this study, virtual screening was used to mine the essential oil components of medicinal plants, and it was found that the components could interact with the spike protein RBD in aerosol to block the interaction of RBD and hACE2, thus cutting off the transmission route and protecting the host. This study has certain guiding significance in the modernization of Traditional Chinese medicine and the prevention of COVID-19.

Amomum tsao-ko is a perennial herbaceous plant of Zingiberaceae, which is a kind of unique Chinese medicinal and edible plant that has been used for thousands of years in China Guo et al., 2017) . In a hot pot, a typical Chinese dish, A.

tsao-ko is an indispensable condiment . It is recorded in the compendium of Materia Medica that A. tsao-ko tastes spicy, warm and belongs to the spleen and stomach meridian Shi et al., 2021; Wei et al., 2021) . A large number of previous studies have shown that A. tsao-ko has multiple pharmacological effects such as sedative, antibacterial and immunity enhancement, and its active substance is mainly volatile essential oil (Qin et al., 2021; Liu et al., 2021; Sabulal and Baby, 2021) . In the early stage of the SARS-CoV-2 outbreak in 2019, both formula 1 proposed by Academician Xiaolin Tong and formula 2 proposed by Academician Qi Wang contained A. tsao-ko (Ai et al., 2020) . Traditional Chinese Medicine (TCM), the treasure of Chinese civilization, is a complex system, which usually acts synergistically (Khanna et al., 2021; Zhao et al., 2021) . It is more suitable to study its mechanism of action by using systematic and holistic research ideas and extracting the complex system from the system for the development of anti-epidemic drugs. Based on the summary of previous studies and the excavation of essential oil components of A. tsaoko, it can be used as a low-cost drug for the development of COVID-19 prevention (Zhu et al., 2021; Lee et al., 2021) .

The COVID-19 pandemic has undergone four major mutations since the outbreak, and the World Health Organization (WHO) began naming in June 2021 after Greek letters: Alpha, Beta, Delta, and the new mutation Omicron that has just been discovered in South Africa (Classification of Omicron (B.1.1.529): SARS-CoV-2 Variant of Concern) (https://www.who.int/news/item/26-11-2021-classification-of-omicron-(b.1.1.529)-sars-cov-2-variant-of-concern; https://www.who.int/en/activities/tracking-SARS-CoV-2-variants/). SARS-CoV-2 is an RNA coronavirus with a simple structure and is prone to mutation. It is mainly composed of four structural proteins, namely spike protein, envelope protein, membrane protein, nucleocapsid protein and sixteen nonstructural proteins. The spike protein consists of S1 and S2, of which S1 is composed of N terminal (NTD, 1~217) and receptor-binding domains (RBD, 319~541) and acts directly on hACE2, which is the first step for the SARS-CoV-2 to contact with the host.

Blocking the binding of the spike protein and hACE2 can protect the host to a great extent. The RBD part of the Omicron mutant has 15 key amino acid mutations, which is the mutant with the most mutations found at present (Fig. 1 ). It has been reported that

Omicron mutations can enhance immune escape, binding ability with receptors, virulence and so on. Therefore, it is of extreme significance to explore a substance that is easier to combine with spike protein RBD in the air, to play a role in epidemic prevention .

In this study, GC/Q-TOF MS was applied to identify and analyze the A. tsao-ko essential oil extracted by hydrodistillation. Meanwhile, homology modeling and molecular dynamics simulation were used to construct and search for key amino acid sites of Omicron's spike protein RBD. The Omicron RBD was docked with the components with a relative area percentage of more than 1% in A. tsao-ko essential oil by molecular docking to demonstrate the relationship between them. The interaction between RBD and hACE2 was inhibited to some extent through the interaction between essential oil compositions and RBD. Since ancient times, spices have been used for epidemic prevention, not to eliminate viruses, but to inhibit the ability of viruses to bind to receptors floating in the atmosphere in poorly ventilated spaces. The purpose of this study is not to develop a drug to cure COVID-19, but to excavate and discover a new substance from natural medicinal plant essential oil that can reduce the binding ability of SARS-CoV-2 to host for COVID-19 epidemic prevention.

Amomum tsao-ko were purchased from Yunnan Province, China and authenticated by Prof. Chunchun Zhang from Zhejiang Chinese Medical University. The dried samples were powdered (0.30 mm) and deposited at 4 ℃ until use. Ultrapure water was manufactured by Milli-Q IQ 7000 purification system (Merck, Germany). Chromatographic grade n-hexane was purchased from Merck (Merck, Germany).

Hydrodistillation was applied for the extraction of A. tsao-ko essential oil (AEO).

with a traditional Clevenger apparatus. An aliquot of 200 g material was introduced into the distillation reaction flask with 2.4 L of water soaking for 30 min before extraction and then extracted for 5 h until the essential oil volume was kept constant.

The essential oil was collected in the receiving flask with a circulating water-cooling system and transferred to an amber-colored vial, dehydrated with anhydrous sodium sulphate. Finally, the essential oil was kept at 4 ℃ with N 2 until being used. The experiment was performed in triplicate.

GC/Q-TOF MS analysis was performed on an Agilent quadrupole time of flight mass spectrometer (Agilent Technology Co., LTD, USA) consisting of an Agilent 7890B gas chromatography and an Agilent 7250 time of flight mass spectrometer, using an HP-5MS capillary column (30 m × 250 μm × 0.25 μm film thickness). The operational conditions of GC/Q-TOF MS analysis were preset as follows: helium was used as the carrier gas at 1.0 mL/min flow rate; split ratio 1:20; injection volume 1 μL; injection temperature 230 ℃; ion source temperature 230 ℃; interface temperature 280 °C; oven temperature progress: 60 ℃ (holding for 2 min), 60-120 ℃ (6 ℃/min, holding for 2 min), 120-180 ℃ (5 ℃/min), 180-280 ℃ (30 ℃/min); ionization mode used at electronic impact 70 eV; mass range 29-500 m/z; TOF-scan mode at 5 spectra/s; solvent delay 4 min. The mass spectra of essential oil compositions in the NIST14 mass spectral library and the fragmentation patterns of the mass spectra with the data published in the literature were applied for the identification of essential oil compositions.

Homology modeling is a highly-accurate method for the prediction and construction of the 3D structure of a protein-based on another protein with a known 3D structure (template). Using the SWISS-MODEL interactive server (Waterhouse et al., 2018) , the 3D structures of Omicron mutant spike protein RBD and NTD domains were constructed based on the structures of wild-type SARS-CoV-2 spike protein RBD (PDB showed that the built structures were of good quality and suitable for further studies.

The built structure of RBD Omicron was analyzed by the Fpocket webserver to predict possible active centres (AC) on the protein surfaces.

Fpocket is an open-source pocket detection package based on Voronoi tessellation and alpha-spheres (Schmidtke et al., 2010) . The PDB file of the RBD Omicron was submitted to Fpocket, and the outputs were analyzed using UCSF Chimera software. Binding sites were the distribution of surrounding residues in AC and acted as the active residues.

Molecular dynamics (MD) simulations were performed using GROMACS 2018.8

on Intel(R) Core (TM) i7-10870H CPU@ 2.20 GHz, using the OPLS-AA/L all-atom force field to treat the protein (Abraham et al., 2015) . RBD was then immersed in a preequilibrated octahedral box of TIP3P water and the system was neutralized. The system was then minimized using the energy gradient convergence criterion set to 0.01 kcal/mol Å2 in four steps involving: (i) an initial 5,000 minimization steps (2,500 with the steepest descent and 2,500 with the conjugate gradient) of only hydrogen atoms, (ii) 20,000 minimization steps (10,000 with the steepest descent and 10,000 with the conjugate gradient) of water and hydrogen atoms, keeping the solute restrained, (iii) 50,000 minimization steps (25,000 with the steepest descent and 25,000 with the conjugate gradient) of protein side chains, water and hydrogen atoms, (iv) 100,000

(50,000 with the steepest descent and 50,000 with the conjugate gradient) of complete minimization. Successively, the water, ions and protein side chains were thermally equilibrated in three steps: (i) 5 ns of NVT equilibration with the Langevin thermostat by gradually heating from 0 K to 300 K, while gradually rescaling solute restraints from a force constant of 10 to 1 kcal/mol Å2, (ii) 5 ns of NPT equilibration at 1 atm with the Berendsen thermostat, gradually rescaling restraints from 1.0 to 0.1 kcal/mol Å2, (iii) 5 ns of NPT equilibration with no restraints. Finally, a production run of 200 ns was performed using a timestep of 2 fs. The SHAKE algorithm was used for those bonds containing hydrogen atoms in conjunction with periodic boundary conditions at constant pressure and temperature, particle mesh Ewald for the treatment of long-range electrostatic interactions, and a cutoff of 10 Å for nonbonded interactions.

The computational molecular docking of hACE2 (PDB ID: 6M17.B, receptor)

with Omicron mutant spike protein RBD (ligand) was explored by an informationdriven flexible docking approach using HADDOCK and Autodock Vina software.

Before docking, the ligand and receptor are dehydrated, hydrogenated, and charged. In docking, the receptor was considered semi-rigid while the ligand was flexible, and the grid matrix of the docking area was sized to ensure complete coverage of the protein molecule. The size of the docking range was set to (40,60,60), and the docking centre was (-32,27,17) . The best Autodock Vina score with the suitable energetically favored conformations was used for further analysis. The interaction of AEO compositions with amino acid residues of omicron mutant spike protein RBD was analyzed and represented using ChimeraX.

The chemical compositions of AEO were determined and listed in Fig. 2 and Table   1 . And twenty components with a relative area percentage of more than 1% were identified in AEO representing 92.82% of the total AEO constituents. The total percentage of monoterpenes was the most abundant compositions in AEO (77.73%), including monoterpene hydrocarbons (14.57%) of α-Phellandrene, β-Pinene, α-Pinene, Linalool, (-)-cis-Sabinol. The oxygenated monoterpene compounds possessed higher dipolar moments in essential oils and interacted more vigorously with heating, and they can be extracted more easily than monoterpene hydrocarbons.

Due to the rich essential oil components, Chinese herbal aromatic plants not only have a special aroma, but also have the effect of medical care that are widely used in many industries. Essential oil can cause aroma movement and regulate qi 1 , and have the functions of regulating emotions, coordinating human physiological functions, calming and improving human immunity. Chinese herbal aromatic essential oil can improve the therapeutic outcome of metabolic syndrome by interfering with insulin signal transduction, resisting oxidative stress, reducing the inflammatory response and increasing insulin sensitivity. These results suggest that Chinese herbal aromatic essential oil has the advantages of high efficiency, low toxicity, multi-target and multilevel therapy, and has broad research prospects in the future . As the predominant component in AEO, 1,8-cineole is an oxygenated monoterpene with a fragrance, camphoraceous features and a spicy flavor. Therapeutic applications of 1,8cineole for the treatment of respiratory diseases, muscle pain, neurosis, rheumatism and kidney stones have been investigated as well (Cai et al., 2021) . As an important part of natural essential oil, traditional Chinese medicine essential oil has a wide variety of sources and has high medicinal, industrial and edible values. (Toelzer et al., 2020) . This pocket has been seen occupied by a lipid similar to linoleic acid in an unusually rigid state of spike protein, where all RBDs are found in a locked-down configuration stabilized by lipid-bridged quaternary interactions between adjacent RBDs, and several were previously shown to modulate ACE2 binding and/or antibody evasion (Harvey et al., 2021) . Thus, understanding the underlying molecular basis for the enhanced transmissibility, immune resistance, and virological properties of the Omicron variant may facilitate halting this looming crisis.

As the Omicron variant showed a considerable number of positively charged amino acids at sites of mutation, the electrostatic potential at the RBD-ACE2 interface and isoelectric point (pI) change of mutation sites on RBD were analyzed. It can be found that the increased pronouncedly electro positivity in the Omicron variant (T478K, Q493R, Q498R and Y505H), and also lost a positively charged amino acid at the interaction interface of K417N (Fig. S2) . Combined with existing studies, the largest difference between RBD WT and RBD Omicron was found at positions 417 and 493 (Socher et al., 2021) . For the Omicron variant that exchanged K417 to N417, a strongly reduced interaction with D30, while Q493 mutated to R493 reduced the interaction with K31, which may be due to electrostatic repulsion between the positively charged R and K.

The pI of protein is the pH value at which its surface is completely charged but its net charge is zero. Also, the amino acid pI of a protein can determine the local electric charge of the protein, which is an important basis for finding targets in drug design (Ren et al., 2021; Xia 2021) . A pI value of more than 7 indicates that the protein or amino acid is alkaline, whereas a value less than 7 indicates that it is acidic. At present, there are few studies on pI of RBD mutation of SARS-CoV-2 spike protein (Krebs et al., 2021) . In this study, it is considered that the change of pI has a great influence on the mutation of SARS-CoV-2, which is a problem neglected by the scientific community at present. The theoretical pI of the RBD WT and RBD Omicron variant were 8.38 and 8.73, indicating that the Omicron variant was expected to have more alkaline pI. As shown in Table 2 , the pI of RBD Omicron mutation sites greater than 7 included N440K, T478K, Q493R, Q498R and Y505H. In addition to steric hindrance, the number of basic amino acids also affects the binding affinity between RBD and hACE2. Studies have found that in the omicron virus multiplication process, supplying excessive basic amino acids is required (Hanai, 2022) . Hence, the transmissible strength of COVID-19 variants depended on the mutated basic amino acids that can enhance the binding affinity with ACE-2.

The instability index of less than 40 indicates that the protein structure is stable.

In our study, the range remained 20.43-28.51, indicating the great stability of RBD WT and RBD Omicron . Aliphatic indexes of 70.31 and 73.21 for RBD WT and RBD Omicron indicated that the protein was temperature stable across a wide temperature range. The greater a protein's aliphatic index, the more thermostable it was. A protein with a low grand average of hydropathicity (GRAVY) value is nonpolar and has a stronger affinity for water, indicating that it is intrinsically hydrophilic. The higher amino acid composition of F, L, I and P in RBD Omicron , when compared with RBD WT , suggested that RBD Omicron included more hydrophobic amino acids. The amino acid composition of RBD Omicron was low in polar amino acids such as Q, S and E, while was high in nonpolar amino acids such as L, F and P. These results indicated that the Omicron variant was more hydrophobic and more easily combined with hACE2, which played an important role in increasing its transmissibility.

The final predicted 3D structure model of RBD Omicron was shown in Fig. 3A .

Although there were many active pocket mining tools, there was no statistically significant difference in structure type and tool performance (Clark et al., 2020) . These tools could provide relatively accurate prediction results for protein active pockets regardless of the presence of ligands. Some researchers used Fpocket as an important tool to analyze the protein-ligand binding pocket, screened the active pocket of methionyl-tRNA synthetase, and proved it to be a natural inhibitor of the lead compound with inhibitory ability in subsequent docking experiments (Rowaiye et al., 2022) . Similar studies have also reported that Fpocket can be used as a well-known tool to mine active pockets and predict specific action targets (active pockets) in both the structure and protein of COVID-19, accurately predict active pockets and provide the necessary basis for the accurate positioning of amino acids (Gervasoni et al., 2020 Topological domain-specific average backbone root mean square deviation (RMSD) value of RBD Omicron was less than that of RBD WT as shown in Fig. S3 . For RBD WT , the structure ensembled from simulation (100 ns) deviated from the backbone of RBD WT by 0.30 Å on average, while the RBD Omicron possessed averaged RMSD value of 0.29 Å, indicating that the mutations only slightly altered the structure of RBD Omicron from the RBD WT . Hence, the mutations in the Omicron variant did not significantly reduce the RBD stability, instead, the hACE2-RBD Omicron complex was even slightly more stable than hACE2-RBD WT (Lupala et al., 2022) . Molecular dynamics was used to calculate the root mean square fluctuations (RMSF) value of amino acids. RMSF means the degree of freedom of each amino acid, which can prove to some extent which amino acid residues are more active in space. The residue fluctuations were analyzed by calculating the RMSF of RBD Omicron and RBD WT (Fig. 3B ). According to the average values of RMSF, the RBD Omicron (0.75 Å) was more rigid than RBD WT (0.80 Å). The reduction of RMSF was more pronounced at the interfacing residues of RBD, also known as the receptor-binding motif (Yi et al., 2020) .

HADDOCK (high ambiguity-driven protein-protein docking) algorithm was applied to perform the restraint docking of WT, Delta and Omicron variants with hACE2 receptor by defining the interface residues as reported previously (Koukos et al., 2021; Khandia et al., 2022) . The docking scores for RBD WT -hACE2 and RBD Delta -hACE2 complex as -49.4 kcal/mol and -64.3 kcal/mol, while the docking score for RBD Omicron -hACE2 complex was -141.4 kcal/mol (Table 3 ). The data showed that the reported mutations have increased the binding ability of RBD Omicron to hACE2 than RBD WT and RBD Delta , which was similar to the results previously reported (Khan et al., 2021) . Furthermore, a comparison of van der waals, electrostatic and desolvation energy values also revealed considerable differences among WT, Delta and Omicron variants. For RBD WT and RBD Delta , the van der waals energies were -18.1 kcal/mol and -24.0 kcal/mol, while RBD Omicron had a van der waals energy of -55.2 kcal/mol. The electrostatic energies for RBD WT , RBD Delta and RBD Omicron were -114.9 kcal/mol, -213.6 kcal/mol and -343.6 kcal/mol, respectively. Both the van der waals energy and electrostatic energy of RBD Omicron were increased, and higher electrostatic energy was considered the primary reason for enhanced binding (Khan et al., 2022) .

The interactions at the interface of the ACE2-RBD complex for WT have been previously reported in the perspectives of both static crystal structures and dynamical conformations Lupala et al., 2021) . Cryo-EM showed that eleven of the fifteen mutations in RBD Omicron (K417N, N440K, G446S, S477N , T478K, E484A, Q493R, G496S, Q498R, N501Y, Y505H) were located at ACE2-RBD interface, consequently changing the electrostatics surface changes at the interface. As can be seen from Fig. 4 the salt bridge E484-K31 and K417-D30 of RBD WT -ACE2 were lost in the Omicron variant, while hydrogen bonds Q493-E35, Q498-K353 and Y505-E37

were enhanced by the Omicron mutations, forming new interactions, such as R498-Y41, R498-Q42 and H505-K353. Also, an additional π-π stacking interaction Y501-Y41 was introduced. The key interactions Y449-D38, Y453-H34, A475-S19, N487-Y83, T500-N330 and T500-D355 in RBD WT -ACE2 were preserved in the Omicron variant. Cryo-EM structural analysis of the RBD Omicron -hACE2 complex showed that new salt bridges and hydrogen bonds were formed at R493, S496 and R498 mutations, which may compensate for other Omicron mutations, such as K417N which is known to reduce the affinity between ACE2 and S protein (Mannar et al., 2022) . The interactions R493-E35

have previously been shown to be responsible for the anchor locking and correct orientation of the RBD Omicron when binding to hACE2. Based on docking studies, the Q493R, N501Y, S371L, S373P, S375F, Q498R and T478K mutations contributed significantly to high binding affinity with hACE2. Analysis revealed that mutations at N440K, T478K and N501Y sites may impart ten times more infectivity to WT (Thakur and Ratho, 2022) . The differences in binding pattern between the WT and the Omicron variant complexes revealed that the key mutations are responsible for enhanced binding and the consequent transmissibility and infectivity. Dejnirattisai et al., 2022 found Omicron variant caused widespread escape from neutralization by serum obtained following infection by a range of SARS-CoV-2 variants, meaning that previously infected individuals will have little protection from infection with Omicron.

Since the outbreak of COVID-19, several large-scale mutations have all been carried out around spike proteins, among which RBD mutation is the most common.

The evolution direction of this virus is dependent on transmission from human to human or from animal to human, through the host immune system to complete the screening (Kupferschmidt, 2021) . In short, the pandemic's weakness has led to the evolution of the SARS-CoV-2, which is centred on spike proteins, particularly the RBD part (Sanjuán and Domingo-Calap, 2016) . Traditional Chinese medicine has been playing an important role in the prevention and treatment of COVID-19 from the early stage to the present. Studies have shown that Chinese medicine components have good inhibitory capacity on both structural and non-structural proteins of SARS-CoV-2. The study of Chinese medicine components by network pharmacology is one of the most widely used methods to explore the inhibition of SARS-CoV-2 binding, infection and replication from Chinese medicine at present . Although this research method has some limitations, it can also provide the best binding skeleton in a specific active centre and provide a good research strategy for drug design and drug screening (Sahlan et al., 2021; Dotolo et al., 2021) . Traditional idea of Traditional Chinese medicine is to prevent and control the spread of diseases before they get sick through systematic regulation, which is highly consistent with modern epidemiological views .

Based on the theory of Traditional Chinese medicine, the research idea of blocking the binding of viruses to receptors is more valuable than the discovery of inhibitors of key enzymes of viruses. Therefore, this study focuses on the study of the herbal aromatic plant Amomum tsao-ko. It can be found that all components in AEO can interact with RBD and NTD by molecular docking the components with a relative area percentage of more than 1% with RBD WT , RBD Delta , RBD Omicron , NTD WT and NTD Omicron . The binding energy between AEO components and RBD Omicron varied from -7.0 kJ/mol to -4.5 kJ/mol. A more negative value of binding energy implies stronger binding efficacy (Kar et al., 2020) . And the binding sites of most AEO components were located at AC2 and AC3 or near the interface. Among them, EO02, 03, 08 and 17 with high relative content and strong binding energy with RBD ( Fig. 5 and Table S1 ).

Meanwhile, RBD is the earliest binding region of COVID-19 with the receptor. It was imperative to research spike protein, especially on how to reduce its binding ability with hACE2 in the epidemic prevention of COVID-19. The binding energy of RBD and hACE2 was between -9.8 and -14.0 kcal/mol (COVID-19 spike-host cell receptor GRP78 binding site prediction), which was a relatively strong binding ability. In current studies, the binding energy between no matter the known drug molecules or the drug molecules obtained by virtual screening cannot reach less than -9.8 kcal/mol, that is, the minimum binding energy of RBD and hACE2 (Shah et al., 2020; Cavasotto and Filippo, 2021) , indicating that it was difficult to design the inhibition of spike protein to inhibit viral infection. Therefore, this study focused on reducing the ability of the virus to spread in the air and contact with the host, which was consistent with the results of relevant literature and could provide proof for this study to a certain extent (Abdelli et al., 2021; Kumar et al., 2020; Yu et al., 2020; Naeem et al., 2020; Xiong et al., 2020) .

Virtual screening showed that most components of AEO could interact with AC2 and AC3 of RBD Omicron , indicating that the interaction between AEO and RBD was a synergistic effect of multiple targets (Fig. 5, Fig. 6 and Fig. S4-12) . Spike protein RBD is the most externally exposed part of the SARS-CoV-2 structure. If RBD contact with AEO molecules in the air, the two can interact directly and form a relatively stable complex, thus protecting the host to a certain extent . The essential oil is volatile itself and can interact with SARS-CoV-2 spike protein RBD in the air to inhibit the binding ability of RBD and hACE2, thus achieving epidemic prevention effect.

This study focused on the Epidemic prevention formula 1 and 2 in China, combined with the previous research on the medicinal aromatic plant, Amomum tsaoko. It was found that there were twenty kinds of AEO with a relative area percentage of more than 1% extracted by hydrodistillation and identified by GC/Q-TOF MS.

Meanwhile, AEO possessed a good diffusion effect in the air because of its volatility.

Given the newly emerged Omicron mutation, RBD Omicron was established by homology modeling in this study. Molecular dynamics simulation and the Fpocket method were used to mine its active centres. RBD Omicron was found to bind hACE2 more strongly than RBD WT and may be a potentially highly pathogenic mutant using HADDOCK.

The molecular docking of 20 components in AEO and RBD Omicron showed that they all had a good interaction, and were distributed in three RBD Omicron active centres, and each AEO component and RBD Omicron had a variety of different active centres with similar binding energy. In the absence of direct evidence at present, this study proves that

Omicron is indeed a more pathogenic mutant of COVID-19, excavates essential oils from Traditional Chinese medicine that can inhibit its binding with receptors, and proposes a prevention method for preventing transmission of SARS-CoV-2 prevention.

Although the world is fighting with the virus, the ever-pervasive COVID-19 continues to evolve, posing a challenge to SARS-CoV-2 epidemic prevention. This study aims to prove from the perspective of simulation that the material basis of the medicinal plant Amomum tsao-ko, which has been reported to have SARS-CoV-2 prevention and control ability, namely its natural essential oil components, has the potential inhibitory ability on COVID-19 especially the Omricon mutants. Starting with the spike protein of the Omricon variant, molecular dynamics and Fpocket were used to dig out the active centres and identify the key amino acid residues. The amino acids

in key parts of RBD were described from two aspects of polarity and isoelectric point. 

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. The active centre on RBD, (B) RMSF values of RBD on wild-type (WT) and Omicron mutant, (C) amino acid sequence of RBD on wild-type (WT) and Omicron.

The interactions between RBD-hACE2 of the Omicron variant. Table 3 Molecular docking studies of hACE2 with RBD WT , RBD Delta , RBD Omicron .

☒ The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

☐The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:

HADDOCK score -49.4 ± 4.5 -64.3 ± 13.0 -141.4 ± 5.5

Van der Waals energy (kcal/mol) -18.1 ± 2.6 -24.0 ± 6.6 -55.2 ± 4.9

Electrostatic energy (kcal/mol) -114.9 ± 16.9 -213.6 ± 37.8 -343.6 ± 49.8

Desolvation energy (kcal/mol) -8.3 ± 1.1 -0.6 ± 2.7 -20.1 ± 2.8 Z-Score -0.2 -1.8 -2.0

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Chemistry of Amomum essential oils

Molecular interaction analysis of Sulawesi propolis compounds with SARS-CoV-2 main protease as preliminary study for COVID-19 drug discovery

Mechanisms of viral mutation

fpocket: online tools for protein ensemble pocket detection and tracking

In silico studies on therapeutic agents for COVID-19: Drug repurposing approach

Identification of in vitro-in vivo components of Caoguo using accelerated solvent extraction combined with gas chromatography-mass spectrometry integrated with network pharmacology on indigestion

Computational decomposition reveals reshaping of the SARS-CoV-2-ACE2 interface among viral variants expressing the N501Y mutation

Research progress on aromatic traditional Chinese medicine essential oil intervening metabolic syndrome

OMICRON (B.1.1.529): A new SARS-CoV-2 variant of concern mounting worldwide fear

Free fatty acid binding pocket in the locked structure of SARS-CoV-2 spike protein

Tracking SARS-CoV-2 variants

Specific epitopes form extensive hydrogen-bonding networks to ensure efficient anti-body binding of SARS-CoV-2: Implications for advanced antibody design

Antibody resistance of SARS-CoV-2 variants B.1.351 and B.1.1.7

Structural and functional basis of SARS-CoV-2 entry by using human ACE2

SWISS-MODEL: homology modelling of protein structures and complexes

Chemical profiling of Huashi Baidu prescription, an effective anti-COVID-19 TCM formula

Domains and functions of spike protein in SARS-Cov-2 in the context of vaccine design

Chinese herbal medicine for coronavirus disease 2019: A systematic review and meta-analysis

Key residues of the receptor binding motif in the spike protein of SARS-CoV-2 that interact with ACE2 and neutralizing antibodies

Computational screening of antagonists against the SARS-CoV-2 (COVID-19) coronavirus by molecular docking

TCM prevention and treatment of epidemic disease based on "Preventing before disease, disease prevention before change, prevention and relapse after recovery

Prevention and treatment of COVID-19 using Traditional Chinese Medicine: A review

Analyzing the potential therapeutic mechanism of Huashi Baidu Decoction on severe COVID-19 through integrating network pharmacological methods

University.

The data that support the findings of this study are available from the corresponding author upon reasonable request.