In silico analysis of metabolic pathways for identification of putative drug targets for Staphylococcus aureus (original) (raw)
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Gene, 2015
Multiple Drug Resistant (MDR) bacteria are no more inhibited by the front line antibiotics due to extreme resistance. Methicillin Resistant Staphylococcus aureus (MRSA) is one of the MDR pathogens notorious for its widespread infection around the world. The high resistance acquired by MRSA needs a serious concern and efforts should be carried out for the discovery of better therapeutics. With this aim, we designed a comparison of the metabolic pathways of the pathogen, MRSA strain 252 (MRSA252) with the human host (i.e., Homo sapiens) by using well-established in silico methods. We identified several metabolic pathways unique to MRSA (i.e., absent in the human host). Furthermore, a subtractive genomics analysis approach was applied for retrieval of proteins only from the unique metabolic pathways. Subsequently, proteins of unique MRSA pathways were compared with the host proteins. As a result, we have shortlisted few unique and essential proteins that could act as drug targets against MRSA. We further assessed the druggability potential of the shortlisted targets by comparing them with the DrugBank Database (DBD). The identified drug targets could be useful for an effective drug discovery phase. We also searched the sequences of unique as well as essential enzymes from MRSA in Protein Data Bank (PDB). We shortlisted at least 12 enzymes for which there was no corresponding deposition in PDB, reflecting that their crystal structures are yet to be solved! We selected Glutamate synthase out of those 12 enzymes owing to its participation in significant metabolic pathways of the pathogen e.g., Alanine, Aspartate, Glutamate and Nitrogen metabolism and its evident suitability as drug target among other MDR bacteria e.g., Mycobacteria. Due to the unavailability of any crystal structure of Glutamate synthase in PDB, we generated the 3D structure by homology modeling. The modeled structure was validated by multiple analysis tools. The active site of Glutamate synthase was identified by not only superimposing the template structure (PDB ID: 1E0A) over each other but also by the Parallel-ProBiS algorithm. The identified active site was further validated by crossdocking the co-crystallized ligand (2-oxoglutaric acid; AKG) of PDB ID: 1LLW. It was concluded that the comparative metabolic in silico analysis together with structure-based methods provides an effective approach for the identification of novel antibiotic targets against MRSA.
Biopolymers & Cell, 2019
Aim. To identify novel putative drug targets of methicillin-resistant S. aureus (MRSA) through subtractive proteome analysis. Methods. Identification of non-homologous proteins in the human proteome, search of MRSA essential genes and evaluation of drug target novelty were performed using a protein BLAST server. Unique metabolic pathways identification was carried out using data and tools from KEGG (Kyoto Encyclopedia of Genes and Genomes). Prediction of sub-cellular proteins localization was performed using combination of PSORT v. 3.0.2, CELLO v. 2.5, iLoc-Gpos, and Pred-Lipo tools. Homology modeling was performed using SWISS-MODEL, Phyre2, I-TASSER web-servers and the MODELLER software. Results. Proteomes of six annotated methicillin-resistant strains : MRSA ATCC BAA-1680, H-EMRSA-15, LA MRSA ST398, MRSA 252, MRSA ST772, UTSW MRSA 55 were initially analyzed. The proteome analysis of the MRSA strains in several consequent steps allowed to identify two molecular targets: diadenylate cyclase and D-alanyl-lipoteichoic acid biosynthesis (DltB) protein which meet the requirements of being essential, membrane-bound, non-homologous to human proteome, involved in unique metabolic pathways and new in terms of not having approved drugs. Using the homology modeling approach, we have built three-dimensional structures of these proteins and predicted their ligand-binding sites. Conclusions. We used classical bioinformatics approaches to identify two molecular targets of MRSA :diadenylate cyclase and DltB which can be used for further rational drug design in order to find novel therapeutic agents for treatment of multidrug resistant staphylococcal infection. K e y w o r d s: molecular drug targets; methicillin-resistant Staphylococcus aureus; MRSA; subtractive proteome analysis.
International Journal of Pharmacy and Pharmaceutical Sciences, 2015
Objective: In the present study an attempt has been made by the use of a computational approach to investigate putative drug targets in Staphylococcus aureus. Methods: In silico comparative analysis of the metabolic pathways between the pathogen and the Bos taurus was carried out. Further detection of bacterial genes that are non homologous to host, but are essential for the survival of the pathogen represents a promising means of identifying novel drug targets. Metabolic pathways were obtained from the metabolic pathway database Kyoto Encyclopedia of Genes and Genomes (KEGG) and were compared to identify unique pathways present only in the pathogen and absent in the host. Results: We have identified total 1930 proteins, which are non homologous to Bos taurus protein sequences and among them 374 enzymes are found to be essential for survival of the S. aureus according to the database of essential genes (DEG) database. Further, 10 proteins were predicted as cytoplasmic and cell wall associated proteins, which could serve as potential drug target candidates. Conclusion: The identified potential drug targets form a platform for further investigation in discovery of novel therapeutic agents against S. aureus.
Bioinformation, 2012
The emergence of multidrug-resistant strain of community-acquired methicillin resistant Staphylococcus aureus (CA-MRSA) strain has highlighted the urgent need for the alternative and effective therapeutic approach to combat the menace of this nosocomial pathogen. In the present work novel potential therapeutic drug targets have been identified through the metabolic pathways analysis. All the gene products involved in different metabolic pathways of CA-MRSA in KEGG database were searched against the proteome of Homo sapiens using the BLASTp program and the threshold of E-value was set to as 0.001. After database searching, 152 putative targets were identified. Among all 152 putative targets, 39 genes encoding for putative targets were identified as the essential genes from the DEG database which are indispensable for the survival of CA-MRSA. After extensive literature review, 7 targets were identified as potential therapeutic drug target. These targets are Fructose-bisphosphate aldolase, Phosphoglyceromutase, Purine nucleoside phosphorylase, Uridylate kinase, Tryptophan synthase subunit beta, Acetate kinase and UDP-N-acetylglucosamine 1-carboxyvinyltransferase. Except Uridylate kinase all the identified targets were involved in more than one metabolic pathways of CA-MRSA which underlines the importance of drug targets. These potential therapeutic drug targets can be exploited for the discovery of novel inhibitors for CA-MRSA using the structure based drug design (SBDD) strategy.
International Journal of Peptide Research and Therapeutics, 2021
Staphylococcus aureus infection is a leading cause of mortality and morbidity in community, hospital and livestock sectors, especially with the widespread emergence of methicillin-resistant S. aureus (MRSA) strains. To identify new drug molecules to treat MRSA patients, we have undertaken to search essential proteins that are indispensable for their survival but nonhomologous to human host proteins. The current study utilizes a subtractive genome and proteome approach to screen the possible therapeutic targets against S. aureus USA300. Bacterial essential genes are obtained from the DEG database and are compared to avoid cross-reactivity with human host genes. In silico analysis shows 198 proteins that may be considered as therapeutic candidates. Depending on their sub-cellular localization, proteins are grouped as either vaccine or drug targets or both. Extracellular proteins such as cell division proteins (Q2FZ91, Q2FZ95), penicillin-binding proteins (Q2FZ94, Q2FYI0) of the bacterial cell wall, phosphoglucomutase (Q2FE11) and lipoteichoic acid synthase (Q2FIS2) are considered as vaccine targets, and their epitopes have been mapped. Altogether, 53 drug targets are identified, which have shown similarity with the drug targets available in the DrugBank database. Predicted drug targets belong to the common metabolic pathways of MRSA, such as fatty acid biosynthesis, folate biosynthesis, peptidoglycan biosynthesis, ribosome, etc. Protein-protein interaction analysis emphasizing peptidoglycan biosynthesis reveals the connection between penicillin-binding proteins, mur-family proteins and FemXAB proteins. In this study, staphylococcal FemA protein (P0A0A5) is subjected to structure-based virtual screening for the drug repurposing approach. There are 20 residues missing in the crystal structure of FemA, and 12 of these residues are located at the catalytic site. The missing residues are modelled, and stereochemistry is checked. FDA approved drugs available in the DrugBank database have been used in virtual screening with FemA in search of potential repurposed molecules. This approach provides us with 10 drugs that may be used in the treatment of methicillin-resistant staphylococcal mediated diseases. AutoDock 4.2 is used for in silico screening and shows a comparable inhibition constant (Ki) for all 10 FDA-approved drugs towards FemA. Most of these drugs are used in the treatment of various cancers, migraines and leukaemia. Protein-drug interaction analysis shows that the drugs mostly interact with hydrophobic residues of FemA. Moreover, Tyr328 and Lys383 contribute largely to hydrogen bondings during interactions. All interacting amino acids that bind to the drugs are part of the active site cavity of FemA.
2016
Vancomycin-resistant Staphylococcus aureus (VRSA) is a Gram-positive, facultative aerobic bacterium which is evolved from the extensive exposure of Vancomycin to Methicillin resistant S. aureus (MRSA) that had become the most common cause of hospital and community-acquired infections. Due to the emergence of different antibiotic resistance strains, there is an exigency to develop novel drug targets to address the provocation of multidrug-resistant bacteria. In this study, in-silico genome subtraction methodology was used to design potential and pathogen specific drug targets against VRSA. Our study divulged 1987 proteins from the proteome of 34,549 proteins, which have no homologues in human genome after sequential analysis through CD-HIT and BLASTp. The high stringency analysis of the remaining proteins against database of essential genes (DEG) resulted in 169 proteins which are essential for S. aureus. Metabolic pathway analysis of human host and pathogen by KAAS at the KEGG server sorted out 19 proteins involved in unique metabolic pathways. 26 human non-homologous membrane-bound essential proteins including 4 which were also involved in unique metabolic pathway were deduced through PSORTb, CELLO v.2.5, ngLOC. Functional classification of uncharacterized proteins through SVMprot derived 7 human non-homologous membrane-bound hypothetical essential proteins. Study of potential drug target against Drug Bank revealed pbpA-penicillin-binding protein 1 and hypothetical protein MQW_01796 as the best drug target candidate. 2D structure was predicted by PRED-TMBB, 3D structure and functional analysis was also performed. Protein–protein interaction network of potential drug target proteins was analyzed by using STRING. The identified drug targets are expected to have great potential for designing novel drugs against VRSA infections and further screening of the compounds against these new targets may result in the discovery of novel therapeutic compounds that can be effective against Vancomycin resistant S. aureus.
opportunistic organism that has emerged as one of the predominant pathogens in community and healthcare- associated infections with limited and less effective options for treatment in the face of a rising trend in the emergence of resistant strains. This fact has necessitated the search for alternative targets for development of new drugs. In this present study, a subtractive genomic (proteome) approach was used to identify potential drug targets in methicillin resistant Staphylococcus aureus using strain 252 (MSRA252). The complete proteome of MSRA 252 obtained from Uniprot database was subjected to CD-hit suite for clustering; NCBI BlastP suite against the human proteome to exclude homologous proteins; and sequence homology with Database of Essential Genes(DEG) to determine the indispensability of the proteins for the bacteria survival. The essential proteins were further analyzed to predict the metabolic pathways they were involved in using KEGG automatic annotation server (KAAS) and their subcellular locations using, Uniprot and PsortB suite subsequently. The sequence sorting, segregation and formatting was carried out using UFS Sequence Analysis Application after each successive step. The study identified 291 essential non homologous proteins to human out of 2640. Further analysis with KAAS revealed that 114 (33 predicted membrane-associated) of the essential non homologous proteins were involved in different metabolic pathways in the organism and 60 of these were implicated in pathways unique to the bacteria relative to human (host). The study revealed a number of putative, essential non homologous protein candidates that could be further explored for the development of alternative treatments and vaccines for methicillin resistant Staphylococcus aureus infections
Staphylococcus aureus is a gram positive bacterium, responsible for both community-acquired and hospital-acquired infection, resulting in a mortality rate of 39%. 43.2% resistance to methicilin and emerging resistance to Fluroquinolone and Oxazolidinone, have evoked the necessity of the establishment of alternative and effective therapeutic approach to treat this bacteria. In this computational study, various database and online software are used to determine some specific targets of Staphylococcus aureus N315 other than those used by Penicillin, Quinolone and Oxazolidinone. For this purpose, among 302 essential proteins, 101 nonhomologous proteins were accrued and 64 proteins which are unique in several metabolic pathways of S. aureus were isolated by using metabolic pathway analysis tools. Furthermore, 7 essentially unique enzymes involved in exclusive metabolic pathways were revealed by this research, which can be potential drug target. Along with these important enzymes, 15 non-homologous proteins located on membrane were identified, which can play a vital role as potential therapeutic targets for the future researchers.
Journal of Bacteriology, 2009
infections, it remains a major hospital-and community-acquired opportunistic pathogen. With the emergence of multidrug-resistant S. aureus strains, there is an urgent need for the discovery of new antimicrobial drug targets in the organism. To this end, we reconstructed the metabolic networks of multidrug-resistant S. aureus strains using genome annotation, functional-pathway analysis, and comparative genomic approaches, followed by flux balance analysis-based in silico single and double gene deletion experiments. We identified 70 single enzymes and 54 pairs of enzymes whose corresponding metabolic reactions are predicted to be unconditionally essential for growth. Of these, 44 single enzymes and 10 enzyme pairs proved to be common to all 13 S. aureus strains, including many that had not been previously identified as being essential for growth by gene deletion experiments in S. aureus. We thus conclude that metabolic reconstruction and in silico analyses of multiple strains of the same bacterial species provide a novel approach for potential antibiotic target identification.