Loading

JSM Biochemistry and Molecular Biology

Structural Characterization and Molecular Docking of Polyamine Transporters in Enterobacter Cloacae

Review Article | Open Access
Article DOI :

  • 1. Sao Francisco University, Rua Waldemar César da Silveira, Brazil
  • 2. Sao Francisco University, Avenida Senador Lacerda Franco, Brazil
  • 3. Sao Francisco University, Avenida São Francisco de Assis, Brazil
+ Show More - Show Less
Corresponding Authors
Aline Sampaio Cremonesi, Avenida Sao Francisco de Assis, Braganca Paulista, Sao Paulo, Brazil, Tel: (+55 11) 2454-8000
ABBREVIATIONS

ABC: ATP Binding Cassette; TMD: Transmembrane domains; NDB: Nucleotide-binding domains; ATP: Adenosine triphosphate; SBP: Substrate-binding protein; TMHMM: Transmembrane Hidden Markov Model; KEGG: Kyoto Encyclopedia of genes and Genomes; BLAST: Basic Local tool Alignment Search Tool; SMART: Simples Modular Architecture Research Tool; AcSpm: Acetyl-spermine; SPD: Spermidine; PUT: Putrescine; RMSD: rootmean-square engineering.

INTRODUCTION

The bacterium of the genus Enterobacter cloacae are gram-negative strains, facultative anaerobes, belonging to Enterobacteriaceae. Saprophytic microorganisms are from the environment, but they are also found in the gastrointestinal microbiota of animals and humans. Outside the gastrointestinal tract, it is shown to be a pathogen responsible for several infections, namely, endocarditis, septic arthritis, osteomyelitis, and skin and soft tissue infections. The possible mechanisms of pathogenicity of E.cloacae are complex with the involvement of a series of virulence factors, whose role in the evolution of the disease is still unknown [1]. Antibiotics are one of the most important interventions to control diseases caused by bacteria, however, their misuse has promoted bacterial resistance, becoming a global health problem, as in the case of E.cloacae that affects patients hospitalized in intensive care [2, 3]. Studies carried out between the years 1998 to 2003 reported three outbreaks with up to 23 systemic infections and a mortality rate of 34%. In the last 15 years, 5% of cases of nosocomial sepsis, 5% of nosocomial pneumonia, and 10% of cases of post-surgical peritonitis caused by E.cloacae have been reported. In addition, several studies point to E.cloacae with resistance to several antibiotics, due to the production of beta-lactamase [1].

Obtaining nutrients from the environment by bacteria, including pathogenic ones, is mainly done through transport through the cell membrane, making use of specialized protein complexes. ATP-Binding Cassette type transporters form a superfamily of membrane proteins that carry out the transport of molecules and substrates across the membrane, through import and export mechanisms [4]. Exporter-type transporters are found from simple prokaryotes to complex eukaryotes, participating in the pathogenesis of bacteria, but are involved in other processes such as toxin export and drug resistance, DNA repair, export of peptides, proteins, polysaccharides, and lipids, in addition to participating in cell division [4-6]. Importer-type transporters are only present in prokaryotic organisms and are primarily responsible for the uptake and absorption of essential nutrient sources of phosphorus, nitrogen, sulfur, and other compounds. The relationship between the uptake system using ABC transporters and the physiological processes demonstrated in different bacteria is evident since the internalization of nutrients and different compounds favors the diversity of intracellular activities involved in bacterial virulence and pathogenesis [7,8]. The basic structure of ABC transporters consists of two transmembrane domains (TMDs), which form a pore across the cell membrane, selectively allowing substrate transport, two highly conserved nucleotide-binding domains (NDBs), which carry out the hydrolysis of the molecule adenosine triphosphate (ATP), providing energy for substrate translocation. Also, transporters of the import system have an extra protein component, located in the periplasm region, with a substratebinding protein (SBP) responsible for specifically capturing and delivering the substrate to the transmembrane domain for its internalization [4,7].

Among all the substrates necessary for the survival of pathogenic bacteria are the polyamines formed by two or more amine fractions along the aliphatic chain. These molecules are involved with the pathogenesis process of bacteria and their intracellular concentration and internalization of the environment [9]. The need to constantly obtain polyamines by the cell is due to the involvement of polyamines in processes such as biofilm formation, cell growth, resistance to oxidative stress, and nitrogen storage [10,11]. Previous studies have described several import ABC transporters for virulence in Escherichia coli and Pseudomonas aeruginosa such as PotABCD and PotFGHI transporters, responsible for transporting spermidine and putrescine, respectively, in E.coli; and SpuDEFGHI, in which SpuD performs putrescine uptake and SpuE promotes spermidine uptake in P.aeruginosa [12,13].

Periplasmic-binding proteins, PotD and PotF, present in the PotABCD and PotFGHI system, present in Enterobacter cloacae, are described as polyamine binders and, given the importance of these molecules for the growth and survival of pathogenic bacteria, the present work aimed to identify and to characterize the proteins that make up the PotABCD and PotFGHI complexes from in silico analyzes and molecular docking, seeking possible ligands that already exist in the literature capable of partially or completely inhibiting the uptake of polyamines by ABC transporters.

MATERIALS AND METHODS

Search and identification of ABC transporters in Enterobacter cloacae

The identification of the proteins that constitute the ABC polyamine transporter of Enterobacter cloacae bacteria was performed using the online database Kyoto Encyclopedia of Genes and Genomes and the Basic Local tool Alignment Search Tool BLAST [14] allowing the identification of homologous proteins. The amino acid sequences of homologous proteins were aligned with those of Enterobacter cloacae using the Clustal Omega [15] program to identify conserved amino acids in different bacterial species.

Transmembrane domain, signal peptide region, and protein domain

The permease proteins of the transporters were identified from the analysis of transmembrane domains performed by the program TMHMM [16]. The software reviews transmembrane regions based on the method of hidden Markov models [17] and uses algorithms to identify existing patterns in a given sequence of amino acids, in this case, the presence of transmembrane helices [18,19]. The SBPs proteins were identified from the presence of signal peptide by the Signal P 6.0 program [20]. The signal peptide is characterized by short amino acid sequences, usually present in the N-terminal region of the protein, being responsible for directing the proteins to the plasmatic membrane [21,22]. Other domains such as highly conserved AAA+ in ATPase proteins [23] were identified from the Simples Modular Architecture Research Tool (SMART) program combined with the Uniprot database [24].

Homology modeling of ABC transporters protein and molecular docking

Homology modeling consists of determining the threedimensional structure of a protein from the alignment of amino acid sequences between the target protein and one or more template proteins of known structure [25,26]. The HHPred program [27], was used to carry out the molecular modeling by homology of the SBPs proteins of the ABC transportation of polyamines from E. cloacae and the generated models were validated by the Ramachandran diagram that tests the quality of the three-dimensional structures since it provides the conformation of the phi (φ) and psi (ψ) torsion angles of the amino acid sequence. These twist angles are defined for each of the amino acid residues [28]. The PyMol program [29] was used for the visualization of the models, prediction of the binding site, virtual sorting, and estimation of binding affinity with the ligands [30].

Using the Auto Dock Vina software [31], molecular docking was performed to evaluate the scoring function used in the protein-ligand fit, predicting the binding mode and affinity of a ligand concerning a protein [32]. Molecular docking is a method capable of identifying possible drug candidates, which uses predictions of binding and affinity between a protein and a ligand. The AutoDock Vina program is based on the Monte Carlo method which presents scoring functions to estimate the free energy in the receptor and an exploratory method to show the positional and conformational space of the ligand on the receptor [32,33]. From the literature data, molecules possible to interact with SBPs and promote transporter inhibition were selected for this analysis.

RESULTS AND DISCUSSION

Enterobacter cloacae feature two complete polyamine uptake transporters

Two polyamine-importing ABC transporters were identified in Enterobacter cloacae bacteria encoded by the potABCD and potFGHI operons (Figure 1). TMHMM analysis identified six transmembrane regions in PotB, PotC, PotH, and PotI proteins, suggesting that these proteins are TMD of ABC transporters. The SignalP 6.0 program identified a probable signal peptide region in the N-terminal region of the PotD and PotF proteins already described as SBPs, and in the PotD protein, the signal peptide is predicted to be located between amino acids 1 to 19 and in the PotF protein between amino acids 1 to 26.

Analysis of the amino acid sequences of the PotA and PotG proteins resulted in the identification of the AAA domain present in the NDB protein of ABC transporters as the P-loop region that contains the alpha-beta-Rossman loop formed by ABC signature nucleotide binding motifs and the Walker A and Walker B regions [35,23] strongly suggesting that PotA and PotG proteins are NDB proteins.

The periplasmic protein-binding pocket conserves amino acids that interact with polyamines

The multiple alignments of SBP protein sequences of polyamine transporters from previously studied bacteria such as Escherichia coli, Klebsiella pneumoniae, Salmonella enterica, Shigella flexneri, Xanthomonas citri and Pseudomonas aeruginosa with the PotD and PotF proteins of E. cloacae, allowed identifying conserved amino acids in the binding pocket of these proteins (Figure 2). The characteristics of the amino acids present in the PotD and PotF binding pocket determine the molecules that interact with these proteins, one of these characteristics being the hydrophobic amino acids. Therefore, the protein-ligand interaction is promoted by the collapse of the organizational structure of water, resulting in entropic gain associated with this disorganization of the system [36]. Such interaction occurs in a specific way, causing a conformational change in the protein, bringing the two globular domains closer together, and changing the protein from an open conformation to a closed one [8,37]. The amino acids identified in the PotD protein that constitute its binding pocket are: W11, T12, E13, Y14, W206, W232 e D234; and the amino acids that make up the PotF protein binding pocket are: W11, S12, D13, Y14, W218, D221, F250 e D252 (Figure 2A). Previous studies carried out in E.coli, P.aeruginosa, and X.citri bacteria, disclosed the presence of conserved amino acids that provide a recognition specificity in the question of the spermidine and putrescine transport, being these, in PotD appearance of a threonine (T12), glutamate (E13) and a tryptophan (W232) and in PotF the presence of serine (S12), aspartate (D13,221,252) and phenylalanine (F250) [13,8] all of them observed in proteins of this study.

For molecular modeling by homology, the following proteins were identified as template proteins for PotD: PotD from Escherichia coli (PDB ID: 1POT), showing 100% sequence identification, and PotD from Streptococcus pneumoniae (PDB ID: 4EQB) with 99.98% identify. PotF proteins from Escherichia coli (PDB ID: 7OYW) served as the template for PotF, presenting 100% identity; Pseudomonas aeruginosa SpuD (PDB ID: 3TTN) with 99.97% identify and Escherichia coli PotF (PDB ID: 1A99) with 89.80% identify. The architecture of the generated templates was observed by the PyMol program (Figure 3A), indicating similarities in the structure of the PotD and PotF proteins, since the conservation of the ancestral structure is crucial for the maintenance of the protein function [38]. This similarity can be observed from the mean square deviation (rootmean-square engineering - RMSD) that compares the folded protein structures, giving the atomic coordinates after an overlap between two structures. The lower the RMSD value, the more similar the detected structures [39]. The aligner between PotD and PotF proteins in PyMol showed an RMSD of 0.656, indicating high similarity between the structures.

Previous studies have shown structural conservation in substrate-binding proteins, and these proteins generally have two globular domains connected through hinge regions which connect the globular domains. Based on the connection regions, it is possible to classify these proteins as type I, II, or III, and type I SBPs have three pleated beta strands in their hinge region. Type II proteins have only two pleated beta strands and type III proteins have an alpha helix connecting the domains [37]. The analysis of the generated models showed that the PotD and PotF proteins present two pleated beta-sheets between the two domains, N and C-terminal, characteristic of type II periplasmic proteins (Figure 3A). The organizational form of folding of polypeptide chains presents the N-terminal and C-terminal region of the polypeptide chain forming domain 1 and the remaining middle region forming domain 2 [37].

Molecular docking identified possible inhibitors for polyamine uptake

Molecular docking indicated a possible interaction between PotD and spermidine, and between PotF and putrescine. The current literature on possible ligands capable of interacting and inhibiting the uptake of polyamines is scarce, but in one study, the cystamine molecule was identified as a competitive inhibitor by nature concerning putrescine, showing itself as a possible inhibitor of its transport [40]. A study carried out with simple conjugations of molecules with spermine, demonstrated that the Acetyl-spermine conjugate (AcSpm) presents a possible ability to inhibit the uptake of polyamines [41]. The best candidates for molecular docking inhibitors were determined from the Gibbs Free Energy of the interaction [42,43]. When carrying out the molecular docking of the PotD protein with the Acetyl-spermine ligand, nine different positions were generated, and three of these positions interacted with the PotD protein binding pocket, with the free energy of -3.3 kcal/mol, -2.8 kcal/mol, and -2.7 kcal/mol (Figure 4A), comparing these values with those obtained from spermidine docking, -3.4 kcal/mol, -3.3 kcal/mol, and -3.2 kcal/ mol (Figure 4B). The docking of the PotF protein also generated nine different positions, of which cystamine showed interaction with the binding pocket in three positions with the free energy of -3.6 kcal/mol, -3.5 kcal/mol, and -3.0 kcal/mol (Figure 4C), in which it was cataloged and compared with the Gibbs free energy values generated from the docking of putrescine, namely -3.9 kcal/mol, -3.6 kcal/mol, and -3.5 kcal/mol (Figure 4D).

Spermine-conjugated molecules can act as a potent inhibitor of polyamine transport in the breast cancer cell line as the Acetylspermine ligand selected in this study for docking with the PotD protein. Acetyl-spermine is a simplified synthetic analog that can bind with equal affinity to the ABC transporter because it is a spermine-conjugated molecule. In this way, it maintains the physicochemical characteristics of polyamines, including hydrophobicity, which enables protein-ligand interaction, inhibiting spermidine uptake [41].

On the other hand, the cystamine molecule is an organic disulfide obtained by the oxidative dimerization of cystamine that presents the connection of a sulfur pair in which the radical ends are interacting with amine groups. This physical-chemical characteristic, in which the sulfur atom is hydrophobic, favorably allows the protein-ligand interaction, since the pocket amino acids are also hydrophobic. Still, in terms of size, cystamine is an inhibitor similar to polyamines, facilitating its entry to interact with the pocket [44].

CONCLUSION

From the results obtained in this research, it is possible to suggest that two transporters import polyamines present in the Enterobacter cloacae bacterium, formed by NDB, TMD, and SBP proteins. The genes encoding these proteins are organized in the form of potABCD and potFGHI operons and are possible spermidine and putrescine transporters, respectively. The PotD and PotF proteins were highly conserved with most of the SBPs described in the literature, presenting two globular domains linked by beta-pleated strands. The binding pocket of both proteins conserved amino acids involved in the interaction with their respective polyamines, suggesting that each one transports a different polyamine, PotD being a possible spermidine scavenger and PotF possibly a putrescine scavenger. These findings were corroborated by molecular docking performed. Still, the cystamine and Acetyl-spermine molecules were identified as possible inhibitors of the PotF and PotD uptake action, respectively, because they present favorable free energy for effective interaction with the binding pockets. These initial data open possibilities for the identification of new molecules that may serve as inhibitors for polyamine uptake in resistant bacteria, thus, more studies will be carried out to identify new inhibitors of the action of ABC transporters in resistant bacteria and to understand their mechanisms of action.

ACKNOWLEDGMENTS

We would like to thank Universidade São Francisco for the scientific initiation program and support in carrying out this project.

REFERENCES
  1. Mezzatesta ML, Gona F, Stefani S. Enterobacter cloacae complex: clinical impact and emerging antibiotic resistance. Future microbiol. 2012; 7: 887-902.
  2. Hutchings MI, Truman AW, Wilkinson B. Antibiotics: past, present and future. Curr Opin Microbiol. 2019; 51:72-80.
  3. Adedeji WA. The treasure called antibiotics. Ann of Ib postgrad med. Ann Ib Postgrad Med. 2016; 14: 56-57.
  4. Marcelo Cassio Barreto de Oliveira, Balan A. The ATP-binding cassette (ABC) transport systems in Mycobacterium tuberculosis: structure, function, and possible targets for Therapeutics. Biology. 2020; 9: 443.
  5. Anthony M, George, Peter M. Jones. Bacterial Membranes: Structural and Molecular Biology. Caister Academic Press. 2014.
  6. Lewis VG, Ween MP, McDevitt CA. The role of ATP-binding cassette transporters in bacterial pathogenicity. Protoplasma. 2012; 249: 919-942.
  7. Locher KP. Mechanistic diversity in ATP-binding cassette (ABC) transporters. Nat Struct Mol Biol. 2016; 23: 487-493.
  8. Cremonesi AS, De la Torre LI, Maximillia Frazão de Souza, Gabriel S Vignoli Muniz, M Teresa Lamy, Cristiano Luis Pinto Oliveira, et al. The citrus plant pathogen Xanthomonas citri has a dual polyaminebinding protein. Biochem Biophys Rep. 2021; 28: 101171.
  9. Phillips MA. Polyamines in protozoan pathogens. J Biol Chem. 2018; 293: 18746-18756.
  10. Michael AJ. Polyamines in eukaryotes, Bacteria, and Archaea. J Biol Chem, 2016; 291: 14896-14903.
  11. Miller-Fleming L, Olin-Sandoval V, Campbell K, Ralser M. Remaining mysteries of molecular biology: The role of polyamines in the cell. J Mol Biol. 2015; 427: 3389-3406.
  12. Krysenko S, Wohlleben W. Polyamine and Ethanolamine Metabolism in Bacteria as an Important Component of Nitrogen Assimilation for Survival and Pathogenicity. Med Sci. 2022; 10: 40.
  13. Donghui Wu, Lim SC, Dong Y, Wu J, Tao F, Zhou L, et al. Structural basis of substrate binding specificity revealed by the crystal structures of polyamine receptors SpuD and SpuE from Pseudomonas aeruginosa. J Mol Biol. 2012; 416: 697-712.
  14. Altschul SF, Madden TL, Schäffer AA, Zhang J, Zhang Z, Miller W, et al. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 1997; 25: 3389-3402.
  15. Chenna R, Sugawara H, Koike T, Lopez R, Gibson TJ, Higgins DG. Multiple sequence alignment with the Clustal series of programs. Nucleic Acids Res, 2003; 31: 3497-3500.
  16. Krogh A, Larsson B, von Heijne G, Sonnhammer EL. Predicting transmembrane protein topology with a hidden Markov model: application to complete genomes. J Mol Biol. 2001; 305: 567-580.
  17. Patra P, Mondal N, Patra BC, Bhattacharya M. Epitope-Based Vaccine Designing of Nocardia asteroides targeting the Virulence Factor MceFamily Protein by Immunoinformatics Approach. Int J Pept Res Ther. 2020; 26: 1165-1176.
  18. Gomes GG. Interação funcional, expressão de genes e filogenia entre proteínas de membrana de Rickettsia spp. e carrapatos. Tese de Doutorado em Bioquímica Agrícola. 2019.
  19. Silva AP. Modelos Ocultos de Markov. Research Gate. 2015.
  20. Armenteros JJA, Tsirigos KD, Sønderby CK, Petersen TN, Winther O, Brunak S, et al. SignalP 5.0 improves signal peptide predictions using deep neural networks. Nat Biotechnol. 2019; 37: 420-423.
  21. Teufel F, Armenteros JJA, Johansen AR, Gíslason MH, Pihl SI, Tsirigos KD, et al. SignalP 6.0 predicts all five types of signal peptides using protein language models. Nat Biotechnol. 2022; 40:1023-1025.
  22. Cocio TA. Caracterização funcional do gene codificador de uma proteína com peptídeo sinal, masA, de Aspergillus fumigatus. 2016.
  23. Puchades C, Sandate CR, Lander GC. The molecular principles governing the activity and functional diversity of AAA+ proteins. Nat Rev Mol Cell Biol. 2020; 21: 43-58.
  24. Letunic I, Bork P. 20 years of the SMART protein domain annotation resource. Nucleic Acids Res. 2018; 46: D493-D496.
  25. Webb B, Sali A. Comparative protein structure modeling using MODELLER. Curr Protoc Bioinformatics. 2016; 54: 5.6.1-5.6.37.
  26. Paulo Henrique Matayoshi Calixto. Aspectos gerais sobre a modelagem comparativa de proteínas. Ciência Equatorial. 2013.
  27. Zimmermann L, Stephens A, Seung-Zin Nam, Rau D, Kübler J, Lozajic M, et al. A completely reimplemented MPI bioinformatics toolkit with a new HHpred server at its core. J Mol Biol. 2018; 430: 2237-2243.
  28. Souza CD, Bessa J, Freitas RD, Oliveira M, Mota K. Estratégia Algorítmica para a Reconstrução e Validação da Estrutura Molecular de Variantes do SARS-CoV-2. Anais do XV Brazilian e-Science Workshop. 2021; 65-72.
  29. DeLano WL. The PyMol Molecular Graphics System. DeLano Scientific. 2002.
  30. Guedes IA, de Magalhães CS, Dardenne LE. Receptor–ligand molecular docking. Biophys Rev. 2014; 6: 75-87.
  31. Trott O, Olson AJ. AutoDock Vina: improving the speed and accuracy of docking with a new scoring function, efficient optimization, and multithreading. J Comput Chem. 2010; 31: 455-461.
  32. Thomas Gaillard. Evolution of AutoDock and AutoDock Vina on the CASF-2013 benchmark. J Chem Inf Model. 2018; 58: 1697-1706.
  33. Fan J, Fu A, Zhang L. Progress in molecular docking. Quantitative Biology. 2019; 7: 83-89.
  34. Soares MR, Facincani AP, Ferreira RM, Moreira LM, de Oliveira JC, Ferro JA, et al. Proteome of the phytopathogen Xanthomonas citri subsp. citri: a global expression profile. Proteome Science. 2010; 8: 1-11.
  35. Pinto AS. Caracterização estrutural e análises funcionais das proteínas periplasmáticas NrtT e PotF de transportadores do tipo ABC de Xanthomonas axonopodis pv. citri. Tese de Doutorado. 2015.
  36. Barreiro EJ, Fraga CAM. Química Medicinal-: As bases moleculares da ação dos fármacos. Artmed Editora. 2014; 3: 10-27.
  37. Rahman MM, Machuca MA, Roujeinikova A. Bioinformatics analysis and biochemical characterisation of ABC transporter-associated periplasmic substrate-binding proteins ModA and MetQ from Helicobacter pylori strain SS1. Biophy Chem. 2021; 272: 106577.
  38. Filho OAS, de Alencastro RB. Modelagem de Proteínas por Homologia. Quimica Nova. 2003; 26: 253-259.
  39. Sargsyan K, Grauffel C, Lim C. How molecular size impacts RMSD applications in molecular dynamics simulations. J Chem Theory Comput. 2017; 13: 1518-1524.
  40. Hoet PH, Dinsdale D, Lewis CP, Verbeken EK, Lauweryns JM, Nemery B. Kinetics and cellular localization of putrescine uptake in human lung tissue. Thorax. 1993; 48: 1235-1241.
  41. Burns MR, Carlson CL, Vanderwerf SM, Ziemer JR, Weeks RS, Cai F, et al. Amino acid/spermine conjugates: polyamine amides as potent spermidine uptake inhibitors. J Med Chem. 2001; 44: 3632-3644.
  42. Junior S, Paulo. Termodinâmica das reações químicas: energia livre de Gibbs e energia química. 2020.
  43. Paula, Reisdorfer F, Fernandes, Mariana Sabo. Estudos de planejamento de ecdisteróides com ação analgésica potencial: docking molecular e de farmacóforo. 2018.
  44. Jeitner TM, Pinto JT, Cooper AJL. Cystamine and cysteamine as inhibitors of transglutaminase activity in vivo. Biosci Rep. 2018; 38: BSR20180691.
Received : 23 May 2023
Accepted : 24 Apr 2023
Published : 16 May 2023
Journals
Annals of Otolaryngology and Rhinology
ISSN : 2379-948X
Launched : 2014
JSM Schizophrenia
Launched : 2016
Journal of Nausea
Launched : 2020
JSM Internal Medicine
Launched : 2016
JSM Hepatitis
Launched : 2016
JSM Oro Facial Surgeries
ISSN : 2578-3211
Launched : 2016
Journal of Human Nutrition and Food Science
ISSN : 2333-6706
Launched : 2013
JSM Regenerative Medicine and Bioengineering
ISSN : 2379-0490
Launched : 2013
JSM Spine
ISSN : 2578-3181
Launched : 2016
Archives of Palliative Care
ISSN : 2573-1165
Launched : 2016
JSM Nutritional Disorders
ISSN : 2578-3203
Launched : 2017
Annals of Neurodegenerative Disorders
ISSN : 2476-2032
Launched : 2016
Journal of Fever
ISSN : 2641-7782
Launched : 2017
JSM Bone Marrow Research
ISSN : 2578-3351
Launched : 2016
JSM Mathematics and Statistics
ISSN : 2578-3173
Launched : 2014
Journal of Autoimmunity and Research
ISSN : 2573-1173
Launched : 2014
JSM Arthritis
ISSN : 2475-9155
Launched : 2016
JSM Head and Neck Cancer-Cases and Reviews
ISSN : 2573-1610
Launched : 2016
JSM General Surgery Cases and Images
ISSN : 2573-1564
Launched : 2016
JSM Anatomy and Physiology
ISSN : 2573-1262
Launched : 2016
JSM Dental Surgery
ISSN : 2573-1548
Launched : 2016
Annals of Emergency Surgery
ISSN : 2573-1017
Launched : 2016
Annals of Mens Health and Wellness
ISSN : 2641-7707
Launched : 2017
Journal of Preventive Medicine and Health Care
ISSN : 2576-0084
Launched : 2018
Journal of Chronic Diseases and Management
ISSN : 2573-1300
Launched : 2016
Annals of Vaccines and Immunization
ISSN : 2378-9379
Launched : 2014
JSM Heart Surgery Cases and Images
ISSN : 2578-3157
Launched : 2016
Annals of Reproductive Medicine and Treatment
ISSN : 2573-1092
Launched : 2016
JSM Brain Science
ISSN : 2573-1289
Launched : 2016
JSM Biomarkers
ISSN : 2578-3815
Launched : 2014
JSM Biology
ISSN : 2475-9392
Launched : 2016
Archives of Stem Cell and Research
ISSN : 2578-3580
Launched : 2014
Annals of Clinical and Medical Microbiology
ISSN : 2578-3629
Launched : 2014
JSM Pediatric Surgery
ISSN : 2578-3149
Launched : 2017
Journal of Memory Disorder and Rehabilitation
ISSN : 2578-319X
Launched : 2016
JSM Tropical Medicine and Research
ISSN : 2578-3165
Launched : 2016
JSM Head and Face Medicine
ISSN : 2578-3793
Launched : 2016
JSM Cardiothoracic Surgery
ISSN : 2573-1297
Launched : 2016
JSM Bone and Joint Diseases
ISSN : 2578-3351
Launched : 2017
JSM Bioavailability and Bioequivalence
ISSN : 2641-7812
Launched : 2017
JSM Atherosclerosis
ISSN : 2573-1270
Launched : 2016
Journal of Genitourinary Disorders
ISSN : 2641-7790
Launched : 2017
Journal of Fractures and Sprains
ISSN : 2578-3831
Launched : 2016
Journal of Autism and Epilepsy
ISSN : 2641-7774
Launched : 2016
Annals of Marine Biology and Research
ISSN : 2573-105X
Launched : 2014
JSM Health Education & Primary Health Care
ISSN : 2578-3777
Launched : 2016
JSM Communication Disorders
ISSN : 2578-3807
Launched : 2016
Annals of Musculoskeletal Disorders
ISSN : 2578-3599
Launched : 2016
Annals of Virology and Research
ISSN : 2573-1122
Launched : 2014
JSM Renal Medicine
ISSN : 2573-1637
Launched : 2016
Journal of Muscle Health
ISSN : 2578-3823
Launched : 2016
JSM Genetics and Genomics
ISSN : 2334-1823
Launched : 2013
JSM Anxiety and Depression
ISSN : 2475-9139
Launched : 2016
Clinical Journal of Heart Diseases
ISSN : 2641-7766
Launched : 2016
Annals of Medicinal Chemistry and Research
ISSN : 2378-9336
Launched : 2014
JSM Pain and Management
ISSN : 2578-3378
Launched : 2016
JSM Women's Health
ISSN : 2578-3696
Launched : 2016
Clinical Research in HIV or AIDS
ISSN : 2374-0094
Launched : 2013
Journal of Endocrinology, Diabetes and Obesity
ISSN : 2333-6692
Launched : 2013
Journal of Substance Abuse and Alcoholism
ISSN : 2373-9363
Launched : 2013
JSM Neurosurgery and Spine
ISSN : 2373-9479
Launched : 2013
Journal of Liver and Clinical Research
ISSN : 2379-0830
Launched : 2014
Journal of Drug Design and Research
ISSN : 2379-089X
Launched : 2014
JSM Clinical Oncology and Research
ISSN : 2373-938X
Launched : 2013
JSM Bioinformatics, Genomics and Proteomics
ISSN : 2576-1102
Launched : 2014
JSM Chemistry
ISSN : 2334-1831
Launched : 2013
Journal of Trauma and Care
ISSN : 2573-1246
Launched : 2014
JSM Surgical Oncology and Research
ISSN : 2578-3688
Launched : 2016
Annals of Food Processing and Preservation
ISSN : 2573-1033
Launched : 2016
Journal of Radiology and Radiation Therapy
ISSN : 2333-7095
Launched : 2013
JSM Physical Medicine and Rehabilitation
ISSN : 2578-3572
Launched : 2016
Annals of Clinical Pathology
ISSN : 2373-9282
Launched : 2013
Annals of Cardiovascular Diseases
ISSN : 2641-7731
Launched : 2016
Journal of Behavior
ISSN : 2576-0076
Launched : 2016
Annals of Clinical and Experimental Metabolism
ISSN : 2572-2492
Launched : 2016
Clinical Research in Infectious Diseases
ISSN : 2379-0636
Launched : 2013
JSM Microbiology
ISSN : 2333-6455
Launched : 2013
Journal of Urology and Research
ISSN : 2379-951X
Launched : 2014
Journal of Family Medicine and Community Health
ISSN : 2379-0547
Launched : 2013
Annals of Pregnancy and Care
ISSN : 2578-336X
Launched : 2017
JSM Cell and Developmental Biology
ISSN : 2379-061X
Launched : 2013
Annals of Aquaculture and Research
ISSN : 2379-0881
Launched : 2014
Clinical Research in Pulmonology
ISSN : 2333-6625
Launched : 2013
Journal of Immunology and Clinical Research
ISSN : 2333-6714
Launched : 2013
Annals of Forensic Research and Analysis
ISSN : 2378-9476
Launched : 2014
Annals of Breast Cancer Research
ISSN : 2641-7685
Launched : 2016
Annals of Gerontology and Geriatric Research
ISSN : 2378-9409
Launched : 2014
Journal of Sleep Medicine and Disorders
ISSN : 2379-0822
Launched : 2014
JSM Burns and Trauma
ISSN : 2475-9406
Launched : 2016
Chemical Engineering and Process Techniques
ISSN : 2333-6633
Launched : 2013
Annals of Clinical Cytology and Pathology
ISSN : 2475-9430
Launched : 2014
JSM Allergy and Asthma
ISSN : 2573-1254
Launched : 2016
Journal of Neurological Disorders and Stroke
ISSN : 2334-2307
Launched : 2013
Annals of Sports Medicine and Research
ISSN : 2379-0571
Launched : 2014
JSM Sexual Medicine
ISSN : 2578-3718
Launched : 2016
Annals of Vascular Medicine and Research
ISSN : 2378-9344
Launched : 2014
JSM Biotechnology and Biomedical Engineering
ISSN : 2333-7117
Launched : 2013
Journal of Hematology and Transfusion
ISSN : 2333-6684
Launched : 2013
JSM Environmental Science and Ecology
ISSN : 2333-7141
Launched : 2013
Journal of Cardiology and Clinical Research
ISSN : 2333-6676
Launched : 2013
JSM Nanotechnology and Nanomedicine
ISSN : 2334-1815
Launched : 2013
Journal of Ear, Nose and Throat Disorders
ISSN : 2475-9473
Launched : 2016
JSM Ophthalmology
ISSN : 2333-6447
Launched : 2013
Journal of Pharmacology and Clinical Toxicology
ISSN : 2333-7079
Launched : 2013
Annals of Psychiatry and Mental Health
ISSN : 2374-0124
Launched : 2013
Medical Journal of Obstetrics and Gynecology
ISSN : 2333-6439
Launched : 2013
Annals of Pediatrics and Child Health
ISSN : 2373-9312
Launched : 2013
JSM Clinical Pharmaceutics
ISSN : 2379-9498
Launched : 2014
JSM Foot and Ankle
ISSN : 2475-9112
Launched : 2016
JSM Alzheimer's Disease and Related Dementia
ISSN : 2378-9565
Launched : 2014
Journal of Addiction Medicine and Therapy
ISSN : 2333-665X
Launched : 2013
Journal of Veterinary Medicine and Research
ISSN : 2378-931X
Launched : 2013
Annals of Public Health and Research
ISSN : 2378-9328
Launched : 2014
Annals of Orthopedics and Rheumatology
ISSN : 2373-9290
Launched : 2013
Journal of Clinical Nephrology and Research
ISSN : 2379-0652
Launched : 2014
Annals of Community Medicine and Practice
ISSN : 2475-9465
Launched : 2014
Annals of Biometrics and Biostatistics
ISSN : 2374-0116
Launched : 2013
JSM Clinical Case Reports
ISSN : 2373-9819
Launched : 2013
Journal of Cancer Biology and Research
ISSN : 2373-9436
Launched : 2013
Journal of Surgery and Transplantation Science
ISSN : 2379-0911
Launched : 2013
Journal of Dermatology and Clinical Research
ISSN : 2373-9371
Launched : 2013
JSM Gastroenterology and Hepatology
ISSN : 2373-9487
Launched : 2013
Annals of Nursing and Practice
ISSN : 2379-9501
Launched : 2014
JSM Dentistry
ISSN : 2333-7133
Launched : 2013
Author Information X