Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar,        K. Kasturi / International J...
Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar,        K. Kasturi / International J...
Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar,        K. Kasturi / International J...
Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar,        K. Kasturi / International J...
Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar,         K. Kasturi / International ...
Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar,        K. Kasturi / International J...
Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar,        K. Kasturi / International J...
Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar,        K. Kasturi / International J...
Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar,        K. Kasturi / International J...
Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar,        K. Kasturi / International J...
Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar,        K. Kasturi / International J...
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  1. 1. Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar, K. Kasturi / International Journal of Engineering Research and Applications (IJERA) ISSN: 2248-9622 www.ijera.com Vol. 3, Issue 1, January-February 2013, pp.023-033 Molecular Modeling and Simulation Studies of Acyl CoA Synthetaseof Mycobacteriumleprae Dhananjay Kumar*, Anshul Sarvate1, Deblina Dey1, Lakshmi Sahitya U2, Kumar Gaurav Shankar3, K. Kasturi2*(Department of Bioinformatics, Dr. D. Y. Patil Biotechnology and Bioinformatics Institute, Pune, Maharashtra 1(Department of Bioinformatics, Dr. D. Y. Patil Biotechnology and Bioinformatics Institute, Pune, Maharashtra). 2(Department of Biotechnology, AcharyaNagarjuna University, Guntur, A. P.) 3(Department of Computer Science, JNU, India)ABSTRACT Leprosy or Hansen’s disease is caused by societies - leprosy was diagnosed conservatively andan obligate intracellular pathogen i.e. thus mostly accurately18. Historically, the bulkyMycobacterium leprae. Leprosy is a numbers of bacterium in the tissues of lepromatousgranulomatous disease of peripheral nerves and patients no doubtfulness led to the aetiologic agent.mucosa of the upper respiratory tract. This Thus, Mycobacteriumleprae, existence suggests thatinfectious disease results in Leprosy reactions it is one of the first bacterium to be determined24.Inthat cause irreversible nerve damage and 1873 first convincing association of adisabilities. The organism requires minimal set of microorganism with a hominian disease, Armauerfunctional genes for its survival. Most of the Hansen27, unconcealed the leprosy bacillus in skingenes are involved in biosynthetic and metabolic biopsies but failed to culture Mycobacteriumleprae.pathways, so the product of these genes can be A century afterwards the nine banded armadillo33,aimed for the novel drug target. Acyl CoA was victimized as a replacement host enabling hugeSynthetase is an enzyme that participates in fatty quantities of the bacillus which has been kept apartacid biosynthesis. The activation of fatty acids by for biochemical and physiological studies59.Acyl-CoA Synthetase is the need of de novo lipid Consequent efforts to corroborate procreation inbiosynthesis, fatty acid catabolism and synthetic media acquired have been equally futile,remodeling of biological membranes. Therefore although metabolic activity can be sensed21.by emphasizing this protein as a drug target can Leprosy is one of the oldest filmed diseases, relic ahelp in the identification of novel drugs to cure serious health problem though prevalence has beenleprosy. A well organized research comprising of low extensively by 1947, as dapsone (4, 4¢-analogue based drug design and molecular diaminodiphenylsulphone) was discovered. At thatdynamics plays a major role in obtaining the lead time it became the only effective, but exclusivemolecules. The bacteria have developed weakly bactericidal, anti-leprosy drug. The figure ofresistance against many of the drugs available in cases of leprosy worldwide remained at roughly 11the market. Therefore identification of the novel million finished to the early-1980s but by then,drug target and potent drug can be helpful in dapsone resistant strains of M. leprae had enlargedbetter prevention of the disease. to appraising levels. The imperative comeback by the World Health Organization to this problem wasKeywords- docking, homology modeling, leprosy, to acquire multi-drug regimens against M. leprae.M.leprae, molecular, molecular dynamics, Since the treatment now included apace antisepticramachandran plot. medicine, rifampicin and also the treatment had good coverage, so the number of cases as expected I Brief Introduction of Leprosy drops down71. At one stage dominance of leprosy Leprosy was originated over 5000 years was around 3 million, though incidence (i.e. rate ofago, almost going back to the Neolithic times50. appearance of new cases) remains as high as beforeThough remaining disfiguring strip conditions were multi-drug therapy was introduced53 andperplexed with leprosy, deformities symptomatic of immunization with BCG34,47, the incidence ofthe disease now illustrious to be caused by disease remains bedevilment with more thanMycobacterium leprae are recognizable in umpteen 690,000 new person reported annnualy72. The mostarchaeological finds24. One of the most famous main usage in the leprosy check in the penultimateexample is the skull of Robert the Bruce that shows millennium has been the launching of multi-drugthe artist symptom in systemic leprosy of nasal therapy (MDT) 54 in 1982; recommendation of theseptum collapse. By gothic times, synchronous WHO study group10. Freely visible long-term multi-archeological relic shows that - at least in few drug therapy that combines rifampicin, clofazimine 23 | P a g e
  2. 2. Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar, K. Kasturi / International Journal of Engineering Research and Applications (IJERA) ISSN: 2248-9622 www.ijera.com Vol. 3, Issue 1, January-February 2013, pp.023-033and dapsone effectively targets the bacterium patch the different drug targets we found a long-chainminimizing the process of drug-resistant strains1. In protein (i.e. Acyl-CoA Synthetase) which is1991 WHO starts to destroy leprosy as a public essential for fatty acid degradation, phospholipidshealth problem which dramatically minimize the remodeling, and the creation of interminable Acyl-global disease prevalence, suggesting a persistent CoA esters that regulates many physiologicalunknown reservoir. In 2006, 259,017 new cases processes60,19. These membrane-bound enzymes actwere reported, out of this 54% of these new reported on non-polar hydrophobic substrates, fatty acids,cases occurred in India, 1 of 118 countries that has generating Acyl-CoA Synthetase, primal reactiveachieved voiding status (downed as figure < 1 intermediates in lipid synthesis pathways that arecase/10,000 assemblage). Brazil, Democratic water-soluble as advantageously as powerfulRepublic of the Congo, Mozambique and Nepal detergents28,65,35,66,37,75. The structure of theseacquire not achieved execution and invoice for 23% membrane proteins has not been solved for theof new cases73. mammalian Acyl CoA Synthetase but homology to The Rational drug use is in this overview a bacterial form, whose structure has beenincidental to the medical therapeutic view received determined, points at peculiar structural features thatat the WHO conference of 1985 in Nairobi2: are consequential for these enzymes acrossRational use of drug requires that forbearing obtain species60,28,61,4,5.medications appropriate to their clinical needs, in In clinical studies, noteworthy advance has beendoses that grapple their own requirements, for a made concerning the immunology andpassable phase of instance and at the smallest outlay immunopathology of leprosy, the genetics of humanto them and their community44. Leprosy was resistance, mechanisms of nerve unhealthiness, andendemic in Norway (amongst a few, stray parts of chemotherapy. In nearly all of these areas,Europe) in Hansens day. Later, it has become more nevertheless, leprosy remains poorly comprehendeddemonstrating in tropical countries, peculiarly - but compared to different leading bacterial diseases14not solely - in poor local societies. Of the 122 and remains a clinically cardinal disease to this day.countries where leprosy was considered disease in1985, 110 score now reached the end of expelling at 1.2. Causesthe state direct by 2003 and leprosy relic a The bodys immune response to thesemipublic upbeat job only in 12 countries22. antigens of the leprosy bacilli may create chances of inflammation in the skin and nerves, known as1.1. Description of the Condition reactions. There are 2 different types of reactions: The bacilli of leprosy are likely spread type 1 reaction or reversal reaction (RR) and type 2through tiny droplets from the nose or mouth from reactions or erythema nodosumleprosum (ENL).infected and raw individuals48. Tissue infected with Reactions may occur during multidrug therapy or itthe leprosy bacilli, M. leprae, contains up to 166 pg can also occur before/after the multidrug therapyof a peculiar phenolic glycolipid for each mg dry wt. and also they are the primary cause of nerve damageof M. Leprae51,11. Leprosy can happen in varied and impairment in leprosy67,76,38. Nerve damageclinical forms, parasitic on the greeting of the vector occurs very slowly and oftentimes it remainsscheme. Some of the persons with a few skin unnoticed or may be it recognizes at a very latepatches and the merchandise of bacilli are relatively stage. So it shows the symptoms of a reaction whichslender; this is classified as paucibacillary (PB) forces people to seek help31,49.leprosy48. Remaining people with many skin patchesand a superior assort of bacilli in their body and are 1.3. Impactclassified as multibacillary (MB) leprosy64,74. Leprosy is most importantly a disablingActually, leprosy is immunologically important 25 disease. The WHO estimates around two to threeand humans are the only famous hosts applicable to million people all over the world because ofthe coefficient of leprosy; so the World Health disabilities to leprosy77. Usually people sufferedOrganization (WHO) currently recommends a 6 and from leprosy, especially because of those visual12-month handling programme for paucibacillary deformities and disabilities, dread discriminationleprosy and multibacillary leprosy, respectively. To and stigmatization. These people may have facedstop such kind of disease novel drug targets are intense social and also some kind of psychologicalrequired in prescript to the organization of new problems29,55,39.drugs against antibacterial tender pathogens.Generally, a target should cater enough selectivity,yielding a drug which is precise or highly selective II Target Identificationagainst the pathogen with respect to the human Leprosy is an unceasing bacterial disease ofhost3. Moreover, the target should be intrinsic for the skin and nerves in the hands and feet and, ingrowing and viability of the pathogen at least under some cases, the lining of the nose62. Leprosy can bethe stipulation of infection 45. While studying about escalating, causing eternal damage to the skin, 24 | P a g e
  3. 3. Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar, K. Kasturi / International Journal of Engineering Research and Applications (IJERA) ISSN: 2248-9622 www.ijera.com Vol. 3, Issue 1, January-February 2013, pp.023-033nerves, limbs, and eyes56. The clinical symptoms of eight 310 helices and 26 ß-strands (Fig.3a,3b). Thisleprosy diversify but most of all it damages the skin, particular protein represents to the family ofnerves, and secretion membranes78. The resultant of adenylate-forming enzymes and also shows thethe muse shows that Acyl-CoA Synthetase protein presence of an A-motif (adenine-binding site;sequence containing 579 amino acid residues with residues 300– 306) and P-motif (phosphate-ID NO (Q9CD78) which plays a vital role in lipid binding site; residues 163–173) which forms themetabolism58. It belongs to ATP dependent AMP- AMP/ATP binding domain, as it is demonstrated bybinding enzyme family. It activates fatty acids, Q-Site Finder41. An additional conserved region ofwhich functions as signaling molecules and are a 25-amino acid long segment, a fatty-acid bindingstructural element of membranes6. It has one AMP region (residues 375–399; FACS signature motif),binding domain (Fig 1)58 since, deletion and decay which is similar to the family of FACS, andof the gene of Acyl-CoA Synthetase causes removal CASTp16 has been used to predict the given bindingof numerous grave metabolic activities12 and it can region.46also be used as a good target to have a good controlof that disease.III Sequence homology and Conserved domain search Acyl-CoA Synthetase protein describes theevolutionary relationship with Long Chain FattyAcid CoA Synthetase40. It has been found that bothare homologous and further the structural propertiesof Fatty Acyl CoA can be used as a reference for thestudy of Acyl CoA. The Pfam65 results demonstrateAMP-binding domain (40-491), which helps, incatalytic commotion of the protein. This partconsists of SER/Thr/Gly colorful area and can beanalyzed again by a conserved Pro-Lys-Gly triplet.(Fig. 1)58 the enzymes family consist of Acetyl CoASynthetase, luciferase and different added intimatelykindred Synthetase58.Fig.1- showing the domain of target protein58 Pfam23 database search revealed one AMP-binding domain. (Fig 2)46 shows multiple sequencealignment of M.tb FadD13 with E. coli,FadDandttLC-FACS, which reveals that there are 3conserved regions: out of them 2 are ATP-AMPbinding domains, residues from 163– 173 arereferred as P-motif, 300–306 called as A-motif andone fatty-acid binding domain, residues 375–399known as FACS signature motif. These domains areconserved within the super family of adenylateforming enzymes. The predicted model for M. Fig.2- Multiple sequence alignment of M. tb adD13tbFadD13consists of 2 domains—a] a large N- with E. coli fadD and ttLC-FACS. The identicalterminal domain (residues 1–395) and b] a small C- residues in the aligned sequences are indicated withterminal domain (402–503) which are further an asterisk (*). P-motif is phosphate-binding siteconnected by a six-amino acid peptide linker, i.e. the colored in blue, A-motif is adenine-binding siteL motif (residues 396– 401). Secondary structure of colored in purple, L-motif is linker motif colored inthe model was analyzed by iMolTalk15, which yellow and fatty acid binding site is indicated indescribes that the structure contains 12 α-helices, green. 46 25 | P a g e
  4. 4. Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar, K. Kasturi / International Journal of Engineering Research and Applications (IJERA) ISSN: 2248-9622 www.ijera.com Vol. 3, Issue 1, January-February 2013, pp.023-033 excellent results achieved experimentally20. This would let the users to carefully use the rapidly generated in-silico protein models in all the contexts where today only experimental structures provide a solid basis: structure-based drug design, analysis of protein function, interactions, antigenic behavior, and rational design of proteins with increased stability or novel functions. In adding together, protein modeling is the merely way to obtain structural information if any how experimental techniques fail, sometimes due to proteins are simply too large for NMR analysis and cannot be crystallized for X-ray diffraction 58. For homology modeling first the target sequence was retrieved from the database (Table1), then BLAST-p was performed against Protein Data Bank (PDB) and the highest scoring entry (high bit score and low e value) was taken as template (Table1). Finally the protein model was generated using modelingsoftwares(Table1). Table-1 – The targets, templates and softwares used for Homology Modeling46,58 Target Seque Temp Softw Reference nce ID late are Acyl Q9CD 1V26 Swiss SuhanyaRama CoA 78 Mode moorthi, S. Synthe (Swiss- ller Venkatesh tase Prot) Fatty CAA1 1ULT Rokk NidhiJatana et. Acyl 6147 y-P al CoA 4.1.SWISS-MODEL (http://swissmodel.expasy.org) is an automated comparative modeling server basically to predict the three dimensional (3D) protein structures32. SWISSMODEL provides several levels of user interaction through its World Wide Web interface: in the ‗first approach mode‘ only an amino acid sequence of a protein is submitted to build a 3D model68. Template selection, alignment and model building are done completely automatedFig.3-Three-dimensional model of M. tb FadD13 a) by the server. The reliability of SWISS-MODEL isSchematic representation of M. tb FadD13. Red continuously evaluated in the EVA-CM project.colour cylinders represent α-helix and blue arrows ROKKY-P57 a server for De novo structurerepresent β-sheets. N and C terminals are prediction by the simfold energy function with therepresented in white colour.46 b) Electrostatic multi-canonical ensemble fragment assembly.potential surface map of the protein with the A- According to the result generated from variousmotif, P-motif and fatty-acid binding site. Positive protein structure evaluation servers, model 3potentials are shown in blue, negative potentials in generated by Rokky-P was found as the best modelred, neutral in white and ligand in green.46 (Table 2)46.IV Homology Modeling The vital aspiration of homology modelingis to predict a structure from its sequence with anaccuracy which will be equivalent to the most 26 | P a g e
  5. 5. Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar, K. Kasturi / International Journal of Engineering Research and Applications (IJERA) ISSN: 2248-9622 www.ijera.com Vol. 3, Issue 1, January-February 2013, pp.023-033 Table No.2 - Quality assessment of the models residue in the most favoured region of obtained by various protein structure prediction Ramachandran plot (Fig. 5)58 when it was evaluated servers46 with a tool called Structure Analysis andStructure PROCHECKa Verify WHAT Verification Server 47 it shows that this very structureprediction 3Db IFc could be used as a good target model for the designserver of drug.58Modeller 65.4% 43.06% -4.822Prime 79.10% 89.27% -3.270SWISS- 83.10% 93.83% -2.121MODELRokky-P- 86.60% 87.50% -0.387Model 1PHYRE 88.3% 87.42% -1.549Rokky-P- 90.50% 76.59% -0.619Model 5Rokky-P- 90.50% 88.69% -0.143Model 2Rokky-P- 90.80% 86.71% -0.143Model 4Rokky-P- 91.50% 88.49% 0.266Model 3 a Percentage of residues in the most favoured region Fig.5- Shows the Ramachandran plot of the protein58 b Percentage of residues having 3D-1D score >0.2 c Ramchandran Z-score, Z-values above 4.0 and Ramachandran plot of the given model describes below -4.0 are very uncommon that 99.8% of the residues lie in the allowed region as shown in Fig. 6 with only 1 residue is available in V Structure Visualization:- disallowed region for the same structure. The As an ensue of Swiss-PDB Viewer, we can VERIFY-3D42 analysis is used to show the predict the 3-dimensional structure of the protein compatibility 3D-1D score >0.2 to be 99.40% Acyl CoA Synthetase(Fig. 4) based on the corresponding to acceptable side chain homologous protein structure Long Chain Fatty environments. ProQ79 also gave a very good Acid CoA Synthetase whose 3-dimensional LGScore of 6.03 and a most importantly MaxSub of structure is already known either with the help of X- 0.17 for the model while ERRAT13 showed the ray Crystallography or NMR. The protein contains overall quality factor to be 79.59% for the model. the AMP binding site as template.58 The ‘what-if quality70 report’ results summarized in (Table 3) indicate that the best sophisticated model showed a Z-score of -2.16 which shows that it is a suitable range for a valid structure. The Z-score of < =-5.0 denotes a poorly refined molecule46. Table 3- What-if quality report (Z-score) for the initial model of FadD13 before performing the MD simulation and for the final model of M. tb FadD13 refined by the MD simulation 46 Backb Backb Side Side Z- one- one- chain- chain score backb side backb -side for one chain one chain all contac contac conta conta conta ts ts ct cts cts Initi −1.25 −3.02 −2.7 −3.3 −3.3 Fig.4- Shows the predicted structure of the protein al Acyl-CoA Synthetase (Red – α helix, Yellow - ß mod Sheet) 58 el Refi −2.24 −0.92 −1.6 −0.8 −2.1VI Evaluation and assessment of ned generated model:- mod The modeled protein which is built on the el basis of 1v26 B-Chain protein describes 83.2% of 27 | P a g e
  6. 6. Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar, K. Kasturi / International Journal of Engineering Research and Applications (IJERA) ISSN: 2248-9622 www.ijera.com Vol. 3, Issue 1, January-February 2013, pp.023-033What-if Fine packing qQualitycControlreport.Average values of the Z-score for all contacts of theprotein can be read as follows: -5.0≤Z-score(guaranteed wrong structure) <−3.0≤Z-score(probably good structure) <−2.0≤Z-score (goodmodel) Fig.7-Blue Colour circled Spots represents the predicted Active Site in the target protein 58 VIII Molecular Docking A drug named as 4 - ((4 - amino 3 chlorophenyl) sulphonyl) phenyl amine (Fig 8) has been generated using the NCI Enhanced Server 26, as an analogue of the first line drug, dapsone and it has also been predicted that the drug have Anti- Myobacterial action which could serve as the ligand. This particular drug was also analyzed by effectuation of Christopher Lipinskis rule-of-five69, which confirms that the designed ligand has the properties and structural features that make molecules much or less like a drug.58Fig.6 – Shows the Ramachandran plot of the finalmodel of the protein 46 Fig.8- Shows the generated ligand 4 - ((4 amino 3-VII Active Site Prediction chlorophenyl) sulphonyl) phenylanmine58 Binding site was characterized by using Q-Site Finder43 and CASTp16 and these were validated The ensue provided by Hex17 shows a quiteby using the information on binding sites in other fine docking between the ligand and the targethomologous proteins.8,30. protein and they are interpreted on the basis ofPutative Active Sites with Spheres, universally binding distance which is measured to be 5.034known as PASS52 is used to predict the active site as Angstrom between the ligand and the active siteshown in the (Fig 7), which could be used as the (Fig 9) with respect to the Tyrosine while Dapsonepossible docking site for the newly developed shows the distance of 6.052 Angstrom with respectligand.58 to the same amino acid. Hence this proves that reduction in distance between the target protein and ligand increases the docking effect between the target protein and its respective ligand.58 28 | P a g e
  7. 7. Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar, K. Kasturi / International Journal of Engineering Research and Applications (IJERA) ISSN: 2248-9622 www.ijera.com Vol. 3, Issue 1, January-February 2013, pp.023-033Fig.9-Showing the docked ligand to the targetprotein58 Many of the substrates like ATP, CoA andvarious fatty acids were docked to M. tbFadD13 bythe help of IFD (Induced Fit Docking) protocol of Fig.10- Docking of multiple ligands (ATP, fatty acidSchrödinger63. ATP and CoA gave the best XP and CoA) to M. tbFadD13 by using induced fitGscore, in terms of kcal/mol. The fatty acids binded docking. A) M. tbFadD13 docked with ATP,to M. tb FadD13 in an order of decreasing binding: lignoceric acid (24:0) and CoA with lignoceric acidcerotic acid>lignoceric acid>palmitic acid>capric shown in pink colour, ATP in purple and CoA inacid according to their scores. M. tb FadD13 has blue. B) Ligplot showing the protein-ligandhigher affinity for very long chain fatty acids interactions in M. tb FadD13 complexed with ATP,especially cerotic (26:0) and lignoceric (24:0) acid lignoceric acid and CoA. ATP is represented by Atpas compared to palmitic (16:0) / capric (10:0) acid 997, lignoceric acid by Faa 998 and CoA by Coaas also observed through experimental studies36. 999.46Docking had also been carried out with otherdifferent ligands in the following order: ATP, fatty IX Molecular Dynamicsacid (lignoceric acid) followed by CoA (Fig. 10a) As the results have obtained from differentand after that the docked complex was further protein structure evaluation servers, out of themrefined using Desmond 2.29. The key amino acids model 3 generated by Rokky-P was preferred as theinteracting with the substrates were identified as: concluding model (Table 2). Desmond 2.09 wasGly166, Lys172, Thr304, Glu305, Thr485,Lys487 forming used for the further molecular dynamics simulationhydrogen bonds with ATP, Tyr362 and Asp371 with of the final model for a period of 12 ns. Framesfatty acid and Thr167, Thr168 ,His170 and Tyr362 with were collected after every 1 ns, energy minimizedCoA as analyzed by LIGPLOT80(Fig. 10b).46 and after that it was evaluated with various protein- evaluation servers. The total energy reaches equilibrium by 10 ns as shown by the stratagem of the total energy versus MD. After a small rearrangement from the initial conformation, the structure is relatively stable during the whole MD as shown by the RMSD map analysis during the 12 ns MD simulation (Fig 11a). Ultimately the final model obtained was evaluated by the ProSA81 program which examines, whether the interaction of each residue with the remainder of the protein is maintained in a favorable manner (Figure 11b) shows that ProSA81 of the desired model gave a Z 29 | P a g e
  8. 8. Dhananjay Kumar, Anshul Sarvate, Deblina Dey, Lakshmi Sahitya U, Kumar Gaurav Shankar, K. Kasturi / International Journal of Engineering Research and Applications (IJERA) ISSN: 2248-9622 www.ijera.com Vol. 3, Issue 1, January-February 2013, pp.023-033score within the acceptable range (-10 to 10, goodProSA scores are negative and depend on lengthof protein). Figure 11c shows that the energyremains negative for almost all amino acid residuesindicating the acceptability of the predicted model.46Fig.11- Analysis of the final model after molecular only chains with less than 1000 residues and a z-dynamics simulation a) RMSD plot of the MD score ≤ 10. The z-score of M. tb FadD13 issimulation as a function of timescale. 46 b) z-plot of highlighted as large dot c) Energy plot of the finalfinal model generated by ProSA. The z-plot shows model obtained by ProSA.46X Conclusion obtained. Finally MD simulation was performed to The protein structure of Acyl CoA refine and validate the generated model. We hopeSynthetase of Mycobacterium leprae is predicted, that the validated model of the protein presented inthis protein can be taken as drug target because it is this study will be a step forward towards theresponsible in fatty acid metabolism. Acyl CoA designing and development of novel drug targetsSynthetase shows homology with Fatty Acyl CoA against leprosy.Synthetase of Mycobacterium tuberculosistherefore we used this protein to assign its REFERENCESstructural and functional properties to our target 1. A. Alter et.al, (2008) Leprosy as a geneticprotein. Both of the proteins were modelled using model for susceptibility to commondifferent softwares and then validated. In infectious disease, Hum GenetRamachandran plot both of them shows almost 2. Amanda Le Grand et al. (1999)Healthsimilar number of residues in the most favoured Policy and Planning; 14(2): 89-102,region. Multiple sequence alignment of FACS of Oxford University Press.M. tuberculosis with E. coli and T. thermophilus 3. AnusuyaShanmugam,shows presence of conserved motifs, namely P- (2010).Bioinformation Journalmotif, L-motif, A-motif and fatty acid binding site; 4. Black PN, DiRusso CC, Metzger AK,this indicates presence of these motifs in Acyl CoA Heimert TL (1992): Cloning, sequencing,Synthetase also. The active site of Acyl CoA and expression of the fadD gene ofSynthetase was predicted and also the derivative of Escherichia coli encoding Acyl coenzymedapsone was generated and was docked with Acyl A Synthetase. J BiolChem,CoA Synthetase, which shows more binding 267(35):25513-25520.affinity of Acyl CoA Synthetase towards the newly 5. Black PN, Zhang Q, Weimar JD, DiRussoformed dapsone derivative as compared to dapsone. CC (1997): Mutational analysis of a fatty Acyl-coenzyme A Synthetase signature The FACS was docked with ATP, CoA motif identifies seven amino acid residuesand various fatty acids. The multiple ligands that modulate fatty acid substratedocking were also done in an order starting from specificity. J Biol Chem. 272(8):4896-ATP, fatty acid (lignoceric acid) and then by CoA, 4903.and as an outcome interacting residues were 30 | P a g e
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