Effect of Azadirachta indica Bio-Compounds against KpsM Protein of Acinetobacter baumannii

Background: Acinetobacter baumannii was considered as a low priority pathogen earlier, and is been now reported as a priority pathogen causing nosocomial infections. Selection of natural compounds to target the organism is the need of the hour. Aim: This study is aimed to target the KpsM protein of A. baumannii with the bio-compounds from Azadirachta indica using in-silico docking analysis. Materials and Methods: KpsM protein was retrieved and optimisation of protein was done. After that optimization and ligand preparation was carried out. It was continued by molinspiration assessment of the molecular properties of selected compounds. It was followed by docking simulation and docking visualisation. Results: Out of the 7 compounds of Azadirachta indica, dihydro diisoeugenol is the best compound to act on the KpsM protein of Acinetobacter baumannii and a binding energy of -6.83Kcal/Mol. Original Research Article Kumaran et al.; JPRI, 33(47B): 773-780, 2021; Article no.JPRI.74430 774 Conclusion: The findings of the study reports isoeugenol with more binding energy than other compounds towards the selected protein KpsM of Acinetobacter baumannii. However it requires further experimental studies to understand the mechanism of its actions and safety.


INTRODUCTION
Acinetobacter baumannii can cause recalcitrant infections of the urinary tract, lungs or in wounds and yet in different components of the body [1]. They additionally colonize or board an affected person without inflicting infections or symptoms, particularly through droplets or open wounds [2,3]. A. baumannii is a growing and foremost problem surviving in dry environments and is dreadful for its fast-spreading nature. It is multidrug resistant and, typically to the last group of drugs, the carbapenems. A.baumannii is a topic of concern for its multidrug-resistant property as it has emerged as a nosocomial pathogen, with the capability to rival MRSA. A.baumannii is categorised under the ESKAPE group of pathogens and belongs to the ACB complex with an extensive property of drug resistance [4].
Many vital virulent genes are attributed for the virulence and resistance property viz., csuE, pgaB, KpsM, epsA, ptk, bfmS and ompA. 98% of the isolates possess more than four biofilm genes. The hydrophobicity profile of KpsM shows that it is a quintessential membrane protein, and is capable of binding to ATP-binding domain. KpsM and KpsT encompass a number of organic processes, such as membrane transport, signal transduction etc. Due to the severity of the multidrug resistance and biofilm formation, most of the antibiotics are unresponsive for the treatment of A. baumannii associated systemic infections [5][6].
In this view, the application of alternative medication would aid in the eradication of A. baumannii. Among many established herbal drugs, Azadirachta indica is moreover a herb that is commonly known as Neem. The medicinal utilities had been defined in particular for neem leaf. Neem leaf and its parts had been tested to exhibit immunomodulatory, anti-inflammatory, antihyperglycemic, antiulcer, antimalarial, antifungal, antibacterial, antiviral, antioxidant, antimutagenic and anticarcinogenic properties. The extract comes from the seeds of the tree and has many diverse conventional uses [7]. The most crucial lively parts are azadirachtin and the others include potent compounds like nimbolinin, nimbin, nimbidin, nimbidol, sodium nimbinate, gedunin, salannin, and quercetin. Neem leaves encompass protein, carbohydrates, minerals, calcium, phosphorus, nutrition C, carotene etc. Drugs from such natural sources can be potent candidates for drug design and the application of bioinformatic tools and servers can be used to evaluate their interactions with the target proteins. The present study was thus undertaken to assess the drug ligand interactions of 7 different compounds from neem with KpsM protein of A.baumannii using computational analysis [8,9].

Study Setting
This was an observational in-silico study done in the Department of Microbiology, Saveetha Dental College and Hospital. Institutional approval for the research was obtained and the SRB approval number is IHEC/SDC/UG-1943/21/150.

Retrieval of KpsM Protein and Optimisation
The crystal shape of KpsM protein turned into received from the RCSB protein records bank. The optimisation of crystal shape of KpsM is performed through the addition of hydrogen atoms. Kollman united atoms pressure subject turned into used to assign digital expenses to the protein atoms which turned into performed withinside the AutoDock device and the RASMOL device turned into used for the visualisation of 3 dimensional shape of KpsM protein.

Ligand Instruction and Optimisation
The systems of the bio-energetic derivatives of A. indica have been received from the chemsketch software. The generated 3-D structures have been then optimised. The decided ligands have passed through next conversions through an open label molecular converter program. Then they have been stored in protein data base (PDB) format.

Molinspiration Evaluation
The counts of hydrogen bond acceptors and donors on the subject of the membrane permeability and bioavailability of the logP for partition coefficient were assessed using the molinspiration evaluation program.

Docking Simulations
The Auto Dock tool was used for docking evaluation to interpret the affinity among biocompounds of A. indica towards the KpsM protein of A. baumannii.

Docking Visualisation
Using Discovery studio visualiser, the hydrogen bond interaction among the bio-compounds of A. indica towards KpsM of A. baumannii have been visualised. Using similarly docking rating assessments, binding affinities, molecular dynamics and electricity simulations, the relative stabilities have been evaluated.

Structural Retrieval of the KpsM Protein from A. baumannii
The sequence of KPSM from Acinetobacter baumannii was retrieved from Uniprot database and its sequence id was SVK37967.1. The structure of KPSM was not available in PDB database. Hence it was modeled using Swissmodel server using the template 6OIH-B Chain. The modeled structure was evaluated using SAVES-PROCHECK server, it was found to be a good quality model as 95.8% of the residue falls in the most favored region and no residues fall in the disallowed region.
The version became fairly viable with 100% series identification with the template. Besides, the Ramachandran plot confirmed 95.8% of residues in maximum favoured areas and without any residues in disallowed areas. The 3d shape of csgA visualized the use of RASMOL with the crimson color denoting the alpha-helix, yellow arrow denoting the beta sheets and white color denoting the turns.

Molinspiration Evaluation Closer to Drug Likeness
Based at the calculation of the ion channel modulation, GPCR ligand, nuclear receptor ligand, kinase inhibitor, enzyme inhibition and protease inhibition, the bioactivity rating prediction of essential additives of A. indica in opposition to KpsM of A. baumannii closer to drug likeness changed into assessed and tabulated (Table 1).

Docking Evaluation of the A. indica Derivatives towards KpsM of A. baumannii
LGA turned into used for choosing the first-class conformers. The bond inter-moves among the vital compounds from A. indica and KpsM of A. baumannii with inside the stick version via a means of discovery studio visualisations among the chosen compounds are shown (Fig. 1). The KpsM protein interactions with biolively compounds from A. indica are shown. The docking scores, quantity of hydrogen bonds formed, torsional strength among the ligands and the medication have been recorded (Table 2).

DISCUSSION
The amazing virulence component of A.
baumannii is its cap potential to shape biofilms in a 4 fundamental step manner via., attachment of micro organism to the surface, formation of micro-colony, maturation of biofilms and compounds for the in-vitro analysis. The oil handled and the oil untreated businesses on assessment confirmed anti-biofilm impact significantly. Biofilm  Selection of the bioactive compounds from A. indica for the present investigation was done based on previous literature. The vital oil compounds from A. indica have been selected for in-silico assessments, because of the hydrophobic biomolecules and can be implemented as nano-formulations. Crude extracts have shown good activity however purification processes need to be done. With the resources of the computational bioinformatics equipment and databases, it is easier to evaluate the drug-ligand interactions. The pose, ligandreceptor properties and the molinspiration assessments hold good for the evaluation of the antimicrobial efficacy of vital compounds from neem.
Comparing the molecular weight of all of the compounds, imidazole-2-carboxylic acid possesses the least molecular weight of 126. bis (2-propyl pentyl) phthalate ester possesses the better molecular weight of 390.36 that correlates with earlier reports . Other compounds confirmed a molecular weight ranging among >200 daltons. Assessments at the hydrogen bonds donor and acceptor property, bis (2-propyl pentyl) phthalate ester had the best wide variety of rotatable bonds of approximately sixteen collectively with the best miLogP cost of 0.17. The TPSA cost (Topological Polar Surface Area) of a compound is a vital evaluation, because it attributes to the oral bioavailabilty of medicine which ought to be <140>.
Evaluation of the general docking energies confirmed that imidazole-2-carboxylic acid were given the best range of hydrogen bonds even as bis(2-propyl pentyl) phthalate has were given the least. Trihydroindole indicates the least binding electricity of -9.39 while bis(2-propyl pentyl) phthalate indicates about -four.07. Dehydrodiisoeugenol possessed a best inhibition consistent while imidazole-2-carboxylic acid confirmed the least inhibition consistent. Ligand efficiency, electrostatic and torsional electricity had been observed to be more in bis (2-propyl pentyl) phthalate ester. We also can infer from the general interplay that imidazole-2-carboxylic acid confirmed 7 hydrogen bond interactions, four van der waals interactions, 1 p -r interplay and 1 p -alkyl interplay which states the stabilization of the binding structures. This changed into accompanied via way of means of 3-Quinolinecarboxylic acid with five hydrogen bonds and bis(2-propyl pentyl) phthalate ester displaying hydrogen bond interplay. Trihydroindole has were given the best Vanderwaal interplay accompanied via way of means of bis (2-propyl pentyl) phthalate. Dihydro diisoeugenol confirmed 2p-r interactions in association with p-alkyl interactions had been observed to be more in dihydroindole. On the opposite hand, handiest 3-Quinoline carboxylic acid confirmed p-sulfur interactions. Dihydro diisoeugenol is considered as the best compound with the binding energy of -6.83. The limitation of the study is that it was conducted as an in-silico based observation. Thus future prospects are set with evaluating the antimicrobial activity using in-vitro and in-vivo studies [12][13][14][15][16][17][18][19][20][21][22][23]. A. baumannii was selected as the study strains as it is one of the predominant nosocomial pathogen with multi-drug resistant property [24,25]. Selection of bio-active compounds from natural sources and the drug ligand interactions in selecting the compounds of drug design is also promising [26,27]. Apart from natural sources it can also be applied for the selection of compounds from marine sources and also for the selection of drug targets [28-32].

CONCLUSION
The findings of the study report dihydro diisoeugenol with more binding energy in comparison to the other compounds towards the selected protein KpsM of Acinetobacter baumannii. The inhibition effect has to be confirmed using the bio-assays further for the drug design and evaluation. So this compound can be considered as the best candidate to design as a novel drug and to treat A. baumannii infections.
However further experimental validation must be done for its efficacy and safety through in-vitro and in-vivo studies.

DISCLAIMER
The products used for this research are commonly and predominantly use products in our area of research and country. There is absolutely no conflict of interest between the authors and producers of the products because we do not intend to use these products as an avenue for any litigation but for the advancement of knowledge. Also, the research was not funded by the producing company rather it was funded by personal efforts of the authors.

FUNDING
The present study was supported by Saveetha Dental College and Hospitals, Saveetha Institute of Medical and Technical Sciences [SIMATS], Saveetha University and Sri Thirumalaipathy Transport.

NOTE
The study highlights the efficacy of "herbal" which is an ancient tradition, used in some parts of India. This ancient concept should be carefully evaluated in the light of modern medical science and can be utilized partially if found suitable.

CONSENT AND ETHICAL APPROVAL
It is not applicable.

COMPETING INTERESTS
Authors have declared that no competing interests exist.