Identification of secondary metabolites from Crescentia cujete as promising antibacterial therapeutics targeting type 2A topoisomerases through molecular dynamics simulation
Introduction
Fluoroquinolones remain one of the most potent bactericidal antibiotics against both Gram-negative and Gram-positive strains [1]. This broad-spectrum activity of the fluoroquinolones has been lucidly demonstrated against the DNA gyrase and topoisomerase IV of bacteria. These enzymes constitute the type IIA topoisomerases (topo2As) essential for chromosomal supercoiling during bacterial DNA synthesis and thus, remain important therapeutic targets for the development of potent fluoroquinolone-like antibacterial drugs [2]. The DNA gyrase (Gyr) is subdivided into GyrA and GyrB, while topoisomerase IV comprises the ParC and ParE subunits [2]. Over the years, new fluoroquinolones have been developed through structural modification of the existing ones [3,4]. While this strategy has yielded more potent drugs with improved antibacterial activity, inherent adverse effects such as dizziness, headache, nausea, risks of mental health, low blood sugar adverse reaction, liver failure, and even death, have undermined their applications in clinical practice [[3], [4], [5]]. Consequently, several fluoroquinolones such as trovafloxacin and grepafloxacin have been minimally used and/or withdrawn from the market [5]. Furthermore, the overuse of the currently available fluoroquinolones such as ciprofloxacin, ofloxacin, and novobiocin, coupled with chromosomal mutations that have resulted in structural alterations in topo2As as potential targets have further limited the use of fluoroquinolones [5,6]. Hence, the identification of new antibacterial agents against topo2As remains imperative. Interestingly, plants and plant-derived compounds have been used in the treatment of human diseases including bacterial infections [7,8]. Hence, exploring plant-derived compounds as novel antibacterial therapeutics could be a viable strategy to identify potent and less toxic drug candidates against bacterial topo2As.
Crescentia cujete L. is an underutilized plant with antibacterial activities against several multidrug-resistant bacteria strains [[9], [10], [11]]. Its ethanolic leaf extract has been demonstrated against Staphylococcus aureus, Bacillus megaterium, Bacillus cereus, Bacillus subtilis, Shigella dysenteriae, and Bacillus cereus with minimum inhibitory concentrations ranging between 2.5 and 4.5 mg/L [12]. Similarly, Parvin et al. [11] investigated the antibacterial activity of the ethanol and chloroform extracts of its leaf and stem bark against S. aureus and Escherichia coli and reported that the chloroform extract was more effective against the tested organisms, with E. coli (29 mm) being the most susceptible at 200 g/disk of the extract. While the antibacterial activities of the different extracts of C. cujete have been attributed to its diverse bioactive metabolites, the associated mechanism of action and the actual metabolite(s) implicated in the process remain elusive and have not been clearly identified [9,12].
The traditional method of drug discovery, which is mainly based on identifying drug candidates using the conventional trial and error screening process, is usually time-consuming and costly with a high risk of failure rate [13]. However, through computational methods such as the use of advanced computer-aided molecular screening tools, lead compounds from plant secondary metabolites with promising antibacterial activities have been identified [14,15]. Also, with better understanding of essential biomolecular processes provided by more advanced computer modeling tools, such as molecular dynamics (MD) simulation, insight on the associated mechanisms of action of therapeutic metabolites have been deciphered [16,17]. Molecular dynamics simulation provides useful information on protein folding, conformational changes, and appreciation of how atoms move at picosecond temporal resolution when ligands interact with bacterial therapeutic targets [16,17]. Hence, in this study, computational methods were adopted to establish the molecular interactions between the lead metabolites from C. cujete and active site amino acid residues of topo2As as antibacterial targets (Fig. 1). Exploring pharmacologically important plants in this manner may help identify potent and less toxic inhibitors of topo2As that could be developed as novel antibacterial therapeutics.
DNA gyrase and topoisomerase IV are found in both Gram-positive and Gram-negative bacteria, with each playing essential roles in replication [2]. More specifically, DNA gyrase (GyrA and GyrB) and topoisomerase IV (ParC and ParE) are the main therapeutic targets in Gram-negative and Gram-positive bacteria, respectively [18,19]. Unfortunately, due to mutations in these four targets/subunits (GyrA and GyrB, ParC and ParE), varying degrees of resistance to fluoroquinolones have been reported [18]. For instance, in S. aureus, a low level of resistance to fluoroquinolones is due to mutations in ParC subunit [19] and the resistance has been shown to increase if the mutation affects both the ParC and GyrA subunits [19]. Therefore, in this study and for the first time, the susceptibility of the topo2A subunits to lead metabolites from C. cujete was undertaken to identify novel drug candidates as potential inhibitors of topo2As.
Section snippets
Identification of C. cujete metabolites
A total of 58 compounds reported to have been isolated from different parts of C. cujete were identified from databases [NPASS (http://bidd.group/NPASS/) and Dr. Duke's Phytochemical and Ethnobotanical Database (https://phytochem.nal.usda.gov/phytochem/search)] and further confirmed from a recently published literature [9].
Collection and preparation of the identified metabolites
The 3D structures of the identified compounds from C. cujete and those of the reference fluoroquinolones [novobiocin with ID 54675769 (standard for GyrB and ParE)], and
Results
The docking scores of the 58 metabolites of C. cujete docked against each of the topo2A subunits are presented in Table S1. Against GyrA, the metabolites had docking scores ranging from −2.9 to −9.5 kcal/mol, with luteolin having the highest score, while against GyrB, the scores ranged between −3.0 and −9.0 kcal/mol, with pinocembrin being the highest ranked compound (Table S1). Similarly, against ParC and ParE, the docking scores were between −3.2 and −8.5 kcal/mol and −3.3 to −7.8 kcal/mol,
Discussion
Plant-derived compounds are one of the viable sources of therapeutics against several debilitating diseases including bacterial infections and have attracted research interest over the years [26]. In this study, promising inhibitors of topo2As were identified from a library of C. cujete metabolites using computational approaches. Initial screening with molecular docking enabled estimation of binding affinity through prediction of ligand interaction and orientation within a protein active site [
Conclusion
This study has identified the secondary metabolites in C. cujete that could elicit their antibacterial effects through inhibition of topo2As as revealed by the affinity, flexibility, compactness, and stability of the resulting complexes of the top five compounds relative to the reference standards. Among the topo2As, higher affinities were observed against GyrA and GyrB relative to ParC and ParE, which suggests that Gram-negative bacteria will be more susceptible to the study metabolites than
Author's contribution
SS conceptualized and supervised the study. JOA, RAA, YD, and KM generated and analyzed the data. JOA, RAA, YD, and KM wrote the manuscript. All authors read and contributed to critical review of the manuscript for intellectual content and approved the submission for publication.
Declaration of competing interest
None to declare.
Acknowledgments
The assistance of the National Research Foundation (NRF- TWAS Doctoral Scholarship, grant number 129950), South Africa, to Mr. J. O. Aribisala is duly and thankfully acknowledged. The Centre for High-Performance Computing (CHPC), South Africa is equally acknowledged for granting access to the computing systems used in this study.
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