Immunoinformatics Study of Procyanidins as Mast Cell Stabilizers

Basu, Sarkar, and Basak: Immunoinformatics Study of Procyanidins as Mast Cell Stabilizers

Authors

INTRODUCTION

Pollen allergens when come to contact human body e.g. skin, respiratory tract, react with antibodies (immunoglobulin type E or IgE) and produce allergic reactions, also known as immediate-type hypersensi-tivity reactions. Allergen bound IgE, which is synthesized in response to allergens during allergic reaction, becomes fixed to specific receptor on the membranes of mast cells and basophils. The high-affinity IgE receptor FcɛRI on mast cells and basophils, is a heteromeric protein containing αßγ2 subunits. Binding of IgE to FcɛRI occurs through direct interactions between Cɛ3 domains of the heavy chains (Fc) of IgE and the extracellular domains of FcɛRIα. α subunit of FcɛRI receptor binds with IgE molecules with high affinity through four surface exposedtryptophan's (W110, W113, W156 and W87).12 This binding plays significant role in initiating intracellular signaling cascades inside mast cell and controls secretion of allergic mediators e.g. histamine.3 A large conformational rearrangement occurs in IgE structure during its binding with high affinity receptor.45 A novel strategy for antiallergic treatment can be postulated by designing molecules that can afliect the receptor binding affinity of IgE.67

Mast cell stabilizer block mast cell degranulation, stabilizing the cell and thereby preventing the release of histamine8 and related mediators e.g. leukotrienes (for basophils) and prostaglandin D2 and IL-5 (for mast cells) during hypersensitivity reaction. Though innumerable synthetic mast cell stabilizers are present9 in our medicinal science, but the search for suitable natural product as mast cell stabilizer is still going on.10 Comparative study shows that higher efficiency for natural product quercetin than synthetic compound cromolyn for inhibiting allergic reactions.11 Several inhibitors of IgEtFcɛRI binding have been identified e.g. an anti-IgE therapeutic antibody (omalizumab) , an engineered protein inhibitor, DARPin E2_799-11, which are used to treat severe allergic asthma.1213Polyphenols which are abundant in apple are proven efliective for allergic rhinitis treatment by preventing degranulation of granulocytes in mast cells.14,15,16 Procyanidins are polymeric form of catechins. In apple, almost 50% to 60% polyphenols are oligomers and polymers of catechins.Dimeric catechins are procyanidin B1, B2, C1 present in apple extract shows antiallergic activity on rhinitis treatment. Among them, procyanidin C1 extracted from unripe apples (Rosaceae mallus) shows anti-allergic effect by various mechanisms e.g. FcɛRI induced degranulation and cytokine production of mast cells. Procyanidin C1 which is related with intra- cellular signalling pathway,prevents FcɛRI-mediated mast cell activation.17 The flow cytometric study is confirmed that polymeric procyanidins supress the binding of IgE antibody to FcɛRI.17 But the molecular mechanism for inhibitory effect on antibody IgE with FcɛRI binding is unknown to all. A compound must have an anti-allergic activity if it inhibits the binding between Ig E antibody and FcɛRI receptor during type I hypersensitivity reaction.

Comparative studies on different procyanidins structures and physico-chemical properties using computational methods will help us to identify apple extract, as herbal medicine in allergic reactions. Computational docking studies are used to elaborate the mechanism of action of various natural products e.g. quercetin glycosides as inhibitor of angiotensin-converting enzyme for the treatment of myocardial infarction18and fatty acids of black cumin oil as inhibitors of P-glycoprotein to improve pharmaceutics of many lifesaving drugs.19

MATERIALS AND METHODS

X-ray crystallographic structures for human IgE, FcɛRIreceptor

The three-dimensional structure of IgE (PDBID 4J4P) and FcɛRI receptor (PDBID 1F2Q) are downloaded from the RCSB protein Data Bank.20

2D and 3D structures of procyanidin B1, B2, C1 and C2

The chemical structures of four procyanidin molecules B1, B2, C1 and C2 are obtained from ZINC database.21 MOL SDF format of these ligands are converted to mol2 file using UCSF Chimera tool.22

Molecular docking study

Molecular docking is a computational method, which forecasts the preferred positioning of two protein molecules e.g.IgE and FcɛRI receptor when bound to each other and form a stable complex using ClusPro 2.2 server.23 Docking study is also used here to investigate the binding affinity of IgE and FcɛRI receptor in presence of four small molecules e.g. procyanidin B1, B2, C1, C2 using SwissDock.24 Binding efficiency of four pro-cyanidin molecules with FcɛRI receptorin absence of IgE molecule with the help of both SwissDock24 and ClusPro 2.223 server is estimated. With the help of lowest binding energy,the procyanidin molecule which act as best mast cell stabilizer, can be identified using the formula-E = 0.40E_{rep} + −0.40E_{att} + 600E_{elec} + 1.00E_{DARS}. Here, repulsive, attractive, electrostatic as well as interactions extracted from the decoys as the reference state, are considered for structure-based pairwise potential calculation in docking.25 Visualization of docking structures with USCF Chimera,22 helps to calculate the distances and torsional angle distortion of binding, with IgE and FcɛRI receptor in presence of four procyanidin molecules. The positions of four surface - exposed tryptophan's (W110, W113, W156 and W87) molecules which are responsible for α subunit of FcɛRI receptor binding with IgE molecules with high affinity, are distorted due to presence of procyanidin molecule. This distortion affects the binding affinity of IgE molecule with its high affinity receptor.

RESULTS

Docking result

α subunit of FcɛRI receptor binds with IgE molecules (Figure 1) with high affinity through four surface - exposed tryptophan's (WHO, WII3, WI56 and W87) (Figure 2) with lowest binding energy −970.3.

Figure 1

Binding of IgE with FcεRI receptor

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Figure 2

Four surface exposed tryptophan's of FcεRI receptor.

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Interactions of procyanidin B1, B2, C1 and C2 with FcɛRI receptor

For procyanidin C1, interacting amino acids are Met 98 and Leu 9 of FcɛRI receptor, which form H bonds with two oxygen atoms along 2.033 Å and 2.027 Å bond lengths. Similarly, procyanidin BI with its O5 atom forms H bond with Met 98 of FcɛRI receptor with bond distance 1.855 Å (Figure 3).

Figure 3

Interactions of procyanidin 81, B2, CI and C2 with FcɛRI receptor.

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Considering binding energy values, it can be concluded that procyanidin C1 molecule stabilises IgE and FcɛRI receptor complex, most efficiently than the other three compounds (-9.053922 Kcal/mole) (Table 1). All the four procyanidin molecules B1, B2, C1 and C2, form stable complexes with high affinity receptor FcɛRI and their binding energies are shown in Table 2. Binding energy of docking compound IgE and FcɛRI receptor is −970.3 where IgE is ligand protein and its receptor is FcɛRI protein. When procyanidin C1binds with FcɛRI receptor binding energy is −811.4 Kcal/mole.

Table 1

Binding energy of IgE with its high affinity receptor in presence and absence of procyanidin molecules.

Docking structuresLowest binding energy
IgE and FcɛRI receptor−970.3 (For ClusPro 2.0)
IgE and FcɛRI receptor and procyanidin B1−8.685186 (For SWISS Dock)
IgE and FcɛRI receptor and procyanidin B2−8.977657 (For SWISS Dock)
IgE and FcɛRI receptor and procyanidin C1−9.053922 (For SWISSDock)
IgE and FcɛRI receptor and procyanidin C2−8.803351 (For SWISSDock)
Table 2

Binding energy of procyanidin molecules with FcɛRI receptor.

Name of compoundsLowest binding energy
For SWISSDock ΔG Kcal/molFor ClusPro 2.0
Procyanidin B1+ FcɛRI−8.715616−953.3
Procyanidin B2 + FcɛRI−9.348962−818.4
Procyanidin C1 + FcɛRI−9.112617−811.4
Procyanidin C2 + FcɛRI−9.834533−854.3

Binding energy calculation

Binding energies of different docking structure obtained from ClusPro 2.0 and SWISSDock servers are shown in Table 1 and 2. Thesedocking structures are compared to hypothesize the effect of IgE molecule on bound structure of procyanidine BI, B2, CI and C2 with FcɛRI receptor.

Conformational change

α subunit of FcɛRI receptor binds with IgE molecules with high affinity through four surface - exposed tryptophans (W110, W113, W156 and W87).When allergen comes in contact with human body , the allergen binds through antigen binding sites of IgE molecule. This allergen bound IgE interacts with its high affinity receptor, which is present on mast cell of human body. In presence of procyanidin C1, a major conformational change occurs in ; subunit of FcɛRI receptor. The distorted structure of receptor protein with Trp 87, Trp 110, Trp 119 and Trp 156, can be visualized in Figure 4.

Figure 4

Positional changes in four surface exposed tryptophan molecules in presence and absence of IgE molecule.

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Binding Procyanidin C1 with FcɛRI receptor after binding with IgE (Model no. 0) shown in blue and binding Procyanidin C1 with FcɛRI receptor in absence of IgE (Model no. 1.1) shown in red. Positional distortion of four tryptophans W87, W110, W113 and W156 shown in second part.

Conformational change in four tryptophan molecules of ; subunit of FcɛRI receptor, when bound with procyanidin C1, in presence and absence of IgE molecule are calculated by bond distance deviation and deviation in torsional angles in Tables 3 and 4.

Table 3

Distance deviation between amino acids TRP 87, TRP 110, TRP156 in two structures.

Procyanidin C1+ FcɛRI + IgE (#0)Procyanidin C1 + FcɛRI (#1.1)Distance
# 0 Trp 87 CH2# 1.1 Trp 87 CH22.535 Å
#0 TRP 156 CZ2#1.1 TRP 156 CZ21.160 Å
# 0 TRP 110 NE1# 1.1 TRP 110 NE10.750 Å
Table 4

Torsional angle change.

Procyanidin C1+ FcɛRI + IgE (#0)Procyanidin C1 + FcɛRI (#1.1)Deviation in Torsion angle
ATOM1ATOM2ATOM3ATOM4
# 0 Trp 87 CE3# 0 Trp 87 CZ3# 1.1 Trp 87 CZ3# 1.1 Trp 87 CE3−0.482
# 0 TRP 110 NE1# 0 TRP 110 CE2# 1.1 TRP 110 NE1# 1.1 TRP 110 CE2−178.286
#0 TRP 156 CD2#0 TRP 156 CE2#1.1 TRP 156 CD2#1.1 TRP 156 CE2−176.830

DISCUSSION

During type I hypersensitivity reaction, allergen binds with IgE molecule. Allergen bound IgE after binding with high affinity FcɛRI receptor, triggers signalling pathway in mast cell. Thus, degranulation reaction starts (release of histamine etc.) from mast cells. Procyanidin molecules which are present in apple in much higher extent, can bind with high affinity receptor of mast cells spontaneously with negative binding energy from molecular docking study. In presence of IgE molecule, which is responsible for mediating allergic reaction, binding energies of procyanidin- FcɛRI complex, are transformed. Not only that, four surface exposed tryptophan residues which are responsible for IgE- FcɛRI interaction, are distorted in their 3D positions in protein, in absence and presence of IgE. This conformational change in procyanidin- FcɛRI complex affects the signal transduction pathway in mast cell degranulation. Thus, procyanidin molecules, specifically procyanidin C1, can inhibit binding of IgE with its high affinity receptor present on biological membrane of mast cell. So, procyanidin molecules can be projected as therapeutic agent (mast cell stabilizers), present in natural resources, in type I hypersensitivity reaction with further experimental verification.

CONCLUSION

This study has shown that procyanidin C1, a polyphenol, present in apple, can be used as natural anti-allergic drug by stabilizing mast cell during type I hypersensitivity reaction after proper experimental verification.

CONFLICT OF INTEREST

There is no conflict of interest

ABBREVIATIONS

IgE

Immunoglobulin E

FcεRI

Fcε receptor I

Trp

Tryptophan amino acid (W)

Pc

Fragment of crystallization

Cε3

Constant region of ε heavy chain domain 3

IL 5

Interleukin 5

DARPins

Designed ankyrin repeat proteins

GRAPHICAL ABSTRACT

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ABOUT AUTHORS

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Anamika Basu: Is an Lecturer in Dept. of Biochemistry, Gurudas College in Kolkata. She received her M.Sc. degree in Biochemistry from Kalyani University and M.Tech in Biotechnology from Jadavpur University. She teaches Immunology, Molecular biology, Cell Biology and Clinical Biochemistry in graduate and post-graduate level. She is a co-author of four book chapters and several journal papers and conference papers. Her research interest includes Computational Biology, Bioinformatics, Molecular modeling, Molecular docking, Molecular Biology and Plant Biology.

SUMMARY

The search for natural antiallergic compounds from plants is an innovative idea for new drug development. Several researchers have been identified various phytochemicals as mast cell stabilizers. Ethnopharmacology study may lead to new bioactive substances for drug design for allergy therapeutics. In our previous study, several natural phytochemicals e.g. flavonoids, stilbenes, coumarins,phenols etc are identified as nutraceuticals in allergic reactions. In Type I hypersensitivityreaction, binding of immunoglobulin E (IgE) with its high affinity receptor (FcɛRI) on mast cell,plays a significant role in allergic reaction. Molecular docking studies provides detailed molecular level interaction between IgE with FcɛRI receptor in presence and absence of four procyanidin molecules, present in apple. Decrease in binding energy and conformational change in ; subunit of FcɛRI receptor occur in presence of procyanidin C1. This conformation change can affect the signal transduction pathway in mast cell degranu-lation. This in-silico study has shown that procyanidins present in apple can act as mast cell stabilizers by changing the binding af^inity of IgE with its receptor FcɛRI during allergic reaction.

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