|Year : 2022 | Volume
| Issue : 3 | Page : 349-352
In silico Study of the interaction of fucoidan with thrombolytic agents
Victoria Evgenievna Suprunchuk
Department of Pharmaceutical Chemistry and Drug Technology, North-Caucasus Federal University, Stavropol, Russia
|Date of Submission||18-May-2022|
|Date of Decision||16-Jun-2022|
|Date of Acceptance||18-Jul-2022|
|Date of Web Publication||17-Sep-2022|
Victoria Evgenievna Suprunchuk
Department of Pharmaceutical Chemistry and Drug Technology, North-Caucasus Federal University, St. Pushkin 1a, Stavropol 355 017
Source of Support: None, Conflict of Interest: None
Background: Tissue plasminogen activator (tPA) is one of the most widely used drugs in thrombolytic therapy. However, due to the inactivation of tPA in the bloodstream and increased risk of bleeding with increasing tPA dosages, the development of targeted delivery systems of tPA is required. For these purposes, it is possible to use fucoidan. The aim of the work was to study the possibility of forming of tPA-fucoidan conjugates and maintaining the activity of the enzyme using molecular docking. Methods: Docking simulations between tPA and fucoidan were performed by use of a docking software AutoDock tools version 1.5.7 and AutoDock 4.2.6. Using “blind docking” to identify the centers of molecular docking approaches of the enzyme (tPA) with the ligand (the active part of the fucoidan structure), as well as to establish the influence of the ligand on the active site of the enzyme. Results: Two “hot spots” of fucoidan binding to the enzyme were found: the region containing SER85-CYS97 residues and the region containing PHE217-TYR223 residues. This interaction can lead to the successful binding of the enzyme and polysaccharide to form a protein-polysaccharide complex. In this case, there may be a lack of suppression of the action of tPA. The interaction with the ligand was found to occur on the surface of the protein molecule. Conclusions: In this study, coupling simulations of interactions of tPA with fucoidan were conducted. The resulting conjugate can be used in the development of systems for the targeted delivery of a thrombolytic agent. This study predicts that the formation of tPA-fucoidan conjugate is a promising approach for optimizing treatment strategies for thrombosis.
Keywords: Conjugate, docking simulations, fucoidan, thrombolytic therapy key, tissue plasminogen activator
|How to cite this article:|
Suprunchuk VE. In silico Study of the interaction of fucoidan with thrombolytic agents. Biomed Biotechnol Res J 2022;6:349-52
| Introduction|| |
The main thrombotic events in medical practice are ischemic stroke, myocardial infarction, pulmonary embolism, and deep venous thrombosis. Fibrinolysis of the clot is carried out using plasmin, which normally exists in the bloodstream in an inactive form (plasminogen). Recombinant forms of tissue plasminogen activators (tPA, EC 188.8.131.52) and thrombolytic agents obtained from bacteria are used as plasminogen activators in medical practice. tPA effectively activates the fibrinolytic system when administered promptly within the “therapeutic window of opportunity,” and is an approved treatment for ischemic stroke. tPA belongs to the serine protease family, and consists of four types of domain: F1, G, kringle, and serine protease domain, where “the F1 and G modules play a major role in the binding of tPA to fibrin.” To enhance the effectiveness of therapy and significantly reduce the therapeutic dose of tPA, it is possible to use combination therapy approaches. Hence, tPA and annexin A2 or minocycline are used together. However, the issue of the safety of using tPA, in particular, preventing hemorrhagic complications, and its inactivation in the bloodstream remains unresolved. Therefore, it is necessary to develop targeted delivery systems of tPA to the site of blood clot formation.
For these purposes, it is possible to use fucoidan. Fucoidan can be applied as a component of the carrier and is currently attracting high attention from academic researchers and pharmaceutical companies. Fucoidan is a branched sulfated heteropolysaccharide composing to 1 → 3, 1 → 4 linked of fucose residues [Figure 1] and having helix-like three-dimensional structure.
Fucoidan has a broad range of biological activities that are highly influenced by its composition and chemical structure. Several works have shown antiviral, anticoagulant, antibiotic, and neuroprotective effects, including therapeutic potential for Alzheimer's disease and others. Of interest is the ability of fucoidan to exhibit a greater affinity for plasminogen activator inhibitor type 1 (PAI-1) than tPA. This phenomenon can be used to protect tPA from PAI-1 when designing a targeted drug delivery system. However, when using particles containing fucoidan as tPA delivery systems, the fibrinolytic activity of the enzyme was reduced by an average of 9% when examined on an agarose plate. The reason for this decrease in activity may be various factors, including the interaction of fucoidan with the active site of the enzyme. Therefore, it is necessary to study the possibility of such an interaction using molecular docking. Analysis of the literature did not reveal such works. The aim of the work was to study the possibility of forming tPA-fucoidan conjugates and maintaining the activity of the enzyme using molecular docking. This study predicts that the formation of tPA-fucoidan conjugates is a promising approach for optimizing treatment strategies for thrombosis.
| Methods|| |
The structure of tPA (P00750) was found in the AlphaFold Protein Structure Database (https://alphafold.ebi.ac.uk/entry/P00750)., The active part of the fucoidan structure was retrieved from PubChem (http://pubchem.ncbi.nlm.nih.gov/search/search.cgi) compound database (SID: 402346915). Docking simulations between tPA and fucoidan were performed by use of a docking software AutoDock tools version 1.5.7 and AutoDock 4.2.6. The best-scoring poses were visually analyzed using BIOVIA Discovery Visualizer version 184.108.40.20698. The tPA molecule was treated as a macromolecule, and the fucoidan fragment was considered a ligand.
| Results|| |
The lowest binding energy of the complex of tPA with fucoidan was established at seven positions. The values of the binding affinity were changed from −4.31 to −5.28 kcal/mol. The results of the interaction of fucoidan with tPA were tabulated in [Table 1].
|Table 1: Indicators of the interaction of fucoidan with tissue plasminogen activator|
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The values of the inhibition constant correspond to the value of the ligand concentration that induces a given response from the target and has a minimum value of 134.59 uM. The best binding mode of these molecules is shown in [Figure 2].
|Figure 2: Best pose of complexed fucoidan with tPA. (a) 3D structure and (b) interaction with amino acids. tPA: Tissue plasminogen activator, 3D: Three-dimensional|
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| Discussion|| |
Fucoidan can form a strong bond with tPA according to high interaction values. In position 5, a greater number of bonds are formed, whose lengths may not exceed 4 Å4. At this position, four such bonds were revealed with CYS86 (3.77 Å), ARG90 (3.87 Å), and CYS91 (3.17 Å), and THR96 (3.41 Å). When interacting with fucoidan on the side of tPA, a response was found most often found from seven enzyme residues: SER85, CYS86, CYS91, THR96, CYS97, PHE217, and ALA222. Moreover, in many cases under consideration, fucoidan forms a hydrogen bond with the residue CYS86, CYS91, and CYS97, which may be associated with the ability of cysteine to form labile hydrogen bonds S-H. O and S-H. S. Hydrogen bonds formed with the participation of SER85, SER498, and GLU498 can be formed due to the oxygen atom of the carboxyl or hydroxyl group, and with the participation of TYR223 due to the hydrogen atom of the hydroxyl group. In addition, a wide network of van der Waals contacts is formed. Conservative residues ARG90, GLY95, THR96, PHE106, LYS117, GLY210, and ARG497 are involved in van der Waals interactions. This makes it possible to identify discrete regions on the surface of the protein that has an affinity for fucoidan. Two “hot spots” of fucoidan binding to the enzyme can be distinguished: a region containing SER85-CYS97 residues and a region containing PHE217-TYR223 residues.
This interaction can lead to the successful binding of the enzyme and the polysaccharide to form a protein–polysaccharide complex. In this case, there is no suppression of the action of tPA. The lack of interactions with tPA protein active site amino acids leads to the saving of the effectiveness of the enzyme. It is known that the manifestation of the activity of the thrombolytic agent is associated with active sites HIS357, ASP406, and SER513 (https://www.uniprot.org/uniprot/P00750). “Blind docking” showed no formation of bonds with Lys429 and Asp477. This indicates that there is no effect on the formation of a salt bridge between these amino acids, which plays an important role in stabilizing the catalytically active conformation. No conformational positions of the ligand influencing the kringle 2 domain a lysyl-binding site of tPA (M28-K33, D55-D57, and N67-W72). The amidolytic activity of single-chain tPA is associated with Lys156. No interaction with this active site has been identified either. The interaction with the ligand was found to occur on the surface of the protein molecule. This will allow the bound fucoidan to directly interact with PAI-1, inhibiting the tPA-PAI-1 complex. However, fucoidan, as a polymer, has a three-dimensional structure that can physically overlap the active site. This provision requires further study and will be considered in further work.
The present docking simulations expand the understanding of the possibility of interaction between the enzyme and the polysaccharide, which allows us to consider fucoidan as a component of tPA-targeted delivery systems.
| Conclusion|| |
Fucoidan can form a strong bond with tPA according to high interaction values. This interaction can lead to successful binding of the enzyme and the polysaccharide to form a protein-polysaccharide complex. The resulting conjugate can be used in the development of systems for targeted delivery of a thrombolytic agent. This study predicts that formation of tPA -fucoidan conjugate is a promising approach for optimizing treatment strategies for thrombosis.
Financial support and sponsorship
The work was carried out within the framework of the Scholarship of the President of the Russian Federation to young scientists and graduate students № -1758.2021.4.
Ethical issue: The study was approved by scientific board of department of Pharmaceutical Chemistry and Drug Technology, North-Caucasus Federal University, Russia.
Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2]