Modeled 3D Structure Research Articles

Purification and characterization of Cc-Lec, C-type lactose-binding lectin: A platelet aggregation and blood-clotting inhibitor from Cerastes cerastes venom

In this study, we reported for the first time the biochemical and structural characterization of Cc-Lec, a C-type lectin purified from Cerastes cerastes venom by affinity chromatography. This lectin was homogeneous by SDS-PAGE, and was shown to be a 34 271.59Da polypeptide by Electrospray mass spectrometry MS-ES-TOF. Its identified sequence of 160 amino acids corresponding to one subunit, revealed a high identity with other related proteins. Cc-Lec modeled 3D structure appeared as homodimer cross-linked by one disulfide bridge. Cc-Lec exhibited a calcium dependent hemagglutinating activity against human group O erythrocytes. Cc-Lec inhibited platelet aggregation induced by ADP, arachidonic acid or fibrinogen suggesting its interaction with their specific receptors namely P2Y1 and/or P2Y12, GPIIb/IIIa and TPα respectively. Cc-Lec was not lethal for mice until 10mg/kg administered by i.p. route. The lectin displayed a lasting anticoagulation on mice plasma even two days post-injection. This anticoagulation seems to be related to its interaction with coagulation factors Xa and IXa. Therefore, Cc-Lec prevented FXa amidolytic activity with Km=4.3310−4μg/mL and ki=14.4μg/mL. It seems to interact with these targets through CRD domain which could make it a good target as a pharmacological promising molecule in thrombosis diagnosis and therapy.

Structural modeling of human organic cation transporters

Human organic cation transporters (hOCTs) belong to solute carriers (SLC) 22 family of membrane proteins that play a central role in transportation of chemotherapeutic drugs for several clinical and pathological conditions, including cancer and diabetes. These transporters mediate drug transport; however, the precise mechanism of drug-binding and transport by them is not fully uncovered yet, partly due to unavailability of any crystal structure record. In this work, we performed a multi-phasic approach to compute the 3D structural models of seven human organic cation transporters (hOCTs) starting from primary protein sequence. Our structure modeling approach included 1) I-TASSER based comparative sequence alignment, threading and ab-initio protein modeling; 2) models comparison with PSIPRED secondary structure prediction; 3) loop modeling for incongruent secondary structure in Chimera 1.10.1; 4) high resolution structure simulation, refinement, energy minimization using ModRefiner, and 5) validation of the structure models using PROCHECK at SAVEs. From structural point, the computed 3D structures of hOCTs consist of a typical major facilitator superfamily (MFS) fold of twelve α-transmembrane helix domains arranged in a manner rendering hOCTs a barrel shaped structure with a large cleft that opens in cytoplasm. The modeled 3D structure of all hOCTs closely resemble to human SLC2A3 (GLUT3) transporter (PDB ID: 5c65) and displayed an outward-open confirmation and putative cyclic C1 protein symmetry. In addition, hOCTs has a large (>100 amino acids) unique extracellular loop between TMH1 and TMH2 having potential glycosylation sites (Asn-Xaa-Ser/Thr) and cysteine residues, both features indicative of putative role in drug binding and uptake. There is an intracellular three/four-helix loop between TMH6 and TMH7 containing putative phosphorylation sites for precise regulation of hOCTs function as drug transporters. There are nine loops of 4 to 11 amino acids length that protrude from membrane, both intracellularly and extracellularly, and connect adjacent TMHs. The 2D structure prediction showed Nin-Cin topology of all hOCTs. In the unavailability of the crystal structures of hOCTs, the 3D structural models computed in-silico and presented herein can be used for studying the mechanism of drug binding and transport by hOCTs.

RESPON SEISMIK STRUKTUR RANGKA DINDING PENGISI YANG DIMODEL DENGAN ELEMEN SHELL PENUH DAN PARSIAL

Abstract: Research on the seismic response of in-filled frame structure has been done with in-filled frame model as full and partial shell elements. The wall is considered active until the maximum load on the full shell models, while the partial shell model using the gradual load with the strength of the wall is considered inactive if the stress of the wall exceeded the wall strength The 4 storey hotel building with full wall in x-direction and wall with opening in y-direction were modeled in SAP 2000 as 3D infilled-frame using full and partial shell element. In Mxy models, both wall were included in the model, while in My models, only the wall in y-direction included. Therefore, 4 models were obtained, there are full shell model MxyShPn and MyShPn and partial shell model MyShPar and MyShPar. In addition, 2 diagonal strut models MxyS and MyS and an open frame model MOF were made as comparison. Prior to model 3D structure, validation models were created using test result condited by other as reference. For that purphose 5 2D models were created there are open frame model MOF, single strut model MST, multiple strut model MSG, full shell model MShPn and partial shell model MShPar. From validation models, it is apparent that the MxyShPar model mimic the behavior of tested structure better than the other models. From the 3D models analysis result show that the displacement in x-direction of MxyShPn, MxyShPar, MxyS were 89%, 85%, 84% smaller than those of MOF, respectively inclusion of wall in the models, also reduce the internal forces and reduse the natural period of the sctructure.

RESPON SEISMIK STRUKTUR RANGKA DINDING PENGISI YANG DIMODEL DENGAN ELEMEN SHELL PENUH DAN PARSIAL

Abstract: Research on the seismic response of in-filled frame structure has been done with in-filled frame model as full and partial shell elements. The wall is considered active until the maximum load on the full shell models, while the partial shell model using the gradual load with the strength of the wall is considered inactive if the stress of the wall exceeded the wall strength The 4 storey hotel building with full wall in x-direction and wall with opening in y-direction were modeled in SAP 2000 as 3D infilled-frame using full and partial shell element. In Mxy models, both wall were included in the model, while in My models, only the wall in y-direction included. Therefore, 4 models were obtained, there are full shell model MxyShPn and MyShPn and partial shell model MyShPar and MyShPar. In addition, 2 diagonal strut models MxyS and MyS and an open frame model MOF were made as comparison. Prior to model 3D structure, validation models were created using test result condited by other as reference. For that purphose 5 2D models were created there are open frame model MOF, single strut model MST, multiple strut model MSG, full shell model MShPn and partial shell model MShPar. From validation models, it is apparent that the MxyShPar model mimic the behavior of tested structure better than the other models. From the 3D models analysis result show that the displacement in x-direction of MxyShPn, MxyShPar, MxyS were 89%, 85%, 84% smaller than those of MOF, respectively inclusion of wall in the models, also reduce the internal forces and reduse the natural period of the sctructure.

Biotechnology and Bioengineering: Volume 113, Number 2, February 2016

Cover Legend Model 3D structure of the catalytic module of recombinant cellobiohydrolase I from Penicillium verruculosum (PvCel7A) with N‐linked glycans: side and bottom view. (Image courtesy of Dr. Alexander Gusakov et al.) image

Protein modeling and molecular dynamics simulation of SlWRKY4 protein cloned from drought tolerant tomato (Solanum habrochaites) line EC520061.

WRKY genes are members of one of the largest families of plant transcription factors and play an important role in response to biotic and abiotic stresses, and overall growth and development. Understanding the interaction of WRKY proteins with other proteins/ligands in plant cells is of utmost importance to develop plants having tolerance to biotic and abiotic stresses. The SlWRKY4 gene was cloned from a drought tolerant wild species of tomato (Solanum habrochaites) and the secondary structure and 3D modeling of this protein were predicted using Schrödinger Suite-Prime. Predicted structures were also subjected to plot against Ramachandran's conformation, and the modeled structure was minimized using Macromodel. Finally, the minimized structure was simulated in the water environment to check the protein stability. The behavior of the modeled structure was well-simulated and analyzed through RMSD and RMSF of the protein. The present work provides the modeled 3D structure of SlWRKY4 that will help in understanding the mechanism of gene regulation by further in silico interaction studies.

Novel Non-Peptide Inhibitors against SmCL1 of Schistosoma mansoni: In Silico Elucidation, Implications and Evaluation via Knowledge Based Drug Discovery.

Schistosomiasis is a major endemic disease known for excessive mortality and morbidity in developing countries. Because praziquantel is the only drug available for its treatment, the risk of drug resistance emphasizes the need to discover new drugs for this disease. Cathepsin SmCL1 is the critical target for drug design due to its essential role in the digestion of host proteins for growth and development of Schistosoma mansoni. Inhibiting the function of SmCL1 could control the wide spread of infections caused by S. mansoni in humans. With this objective, a homology modeling approach was used to obtain theoretical three-dimensional (3D) structure of SmCL1. In order to find the potential inhibitors of SmCL1, a plethora of in silico techniques were employed to screen non-peptide inhibitors against SmCL1 via structure-based drug discovery protocol. Receiver operating characteristic (ROC) curve analysis and molecular dynamics (MD) simulation were performed on the results of docked protein-ligand complexes to identify top ranking molecules against the modelled 3D structure of SmCL1. MD simulation results suggest the phytochemical Simalikalactone-D as a potential lead against SmCL1, whose pharmacophore model may be useful for future screening of potential drug molecules. To conclude, this is the first report to discuss the virtual screening of non-peptide inhibitors against SmCL1 of S. mansoni, with significant therapeutic potential. Results presented herein provide a valuable contribution to identify the significant leads and further derivatize them to suitable drug candidates for antischistosomal therapy.

Similarity-based modeling applied to signal detection in pharmacovigilance.

One of the main objectives in pharmacovigilance is the detection of adverse drug events (ADEs) through mining of healthcare databases, such as electronic health records or administrative claims data. Although different approaches have been shown to be of great value, research is still focusing on the enhancement of signal detection to gain efficiency in further assessment and follow-up. We applied similarity-based modeling techniques, using 2D and 3D molecular structure, ADE, target, and ATC (anatomical therapeutic chemical) similarity measures, to the candidate associations selected previously in a medication-wide association study for four ADE outcomes. Our results showed an improvement in the precision when we ranked the subset of ADE candidates using similarity scorings. This method is simple, useful to strengthen or prioritize signals generated from healthcare databases, and facilitates ADE detection through the identification of the most similar drugs for which ADE information is available.

Homcos : A Server to Search and Model 3D Structures of Protein-Protein and Compound-Protein Complexes

3D complex structures of proteins and other molecules provide a clue for mechanism of interaction. However, few servers for searching complex structures are available. We are developing an updated server HOMCOS (HOmology Modeling of Complex Structure) with more services than the previous version. It separates component molecules of PDB files of complexes, such as proteins, nucleic acids, small chemical compounds, stores their binding relationships. It searches these molecules by BLAST and our chemical structure comparison program dkcombu. Based on found similar complexes, simple template-based models of the complex can be generated by replacing the template with the query amino acid sequence. The service ‘Modeling Homo Protein Multimer’ searches homologous protein complexes with one query protein sequence, whereas the service ‘Modeling Hetero Protein Multimer’ searches with two sequences. The service ‘Modeling Compound-Protein Complexes’ searches with one sequence and one chemical structure. For modeling a bound conformation of a target compound, our flexible overlay program fkcombu is performed to superimpose the target compound onto the template compound bound to a protein. The program transforms a pose and torsion angles of the target molecule to superimpose atom pairs of the target and template molecule determined by 2D-MCS. The service ‘Searching Contacting Molecules for Query Protein’ searches contacting molecules with homologous proteins to a given query sequence, and provide putative interacting molecules with putative binding sites. It is useful to estimate mutational effects on molecular interactions. The service ‘Searching Contacting Proteins for Query Compound’ searching contacting proteins with similar compounds to a given query chemical compound. It may be useful to predict unintended interacting proteins, which cause side effects of the compound. We hope this server may contribute to the platform for drug discoveries and life sciences.

Identification of Novel Rab27a/Melanophilin Blockers by Pharmacophore-Based Virtual Screening

Melanocytes are unique cells that produce specific melanin-containing intracellular organelles called melanosomes. Melanosomes are transported from the perinuclear area of melanocytes toward the plasma membrane as they become more melanized in order to increase skin pigmentation. In this vesicular trafficking of melanosomes, Rab27a, melanophilin, and myosin Va play crucial roles in linking melanosomes to actin-based motors. To identify novel compounds to inhibit binding interface between Rab27a and melanophilin, a pharmacophore model was built based on a modeled 3D structure of the protein complex that describes the essential binding residues in the intermolecular interaction. A pharmacophore model was employed to screen a chemical library database. Finally, 25 virtual hits were selected for biological evaluations. The biological activities of 11 analogues were evaluated in a second assay. Two compounds were identified as having concentration-dependent inhibitory activity. By analyzing structure-activity relationships of derivatives of BMD-20, two hydroxyl functional groups were found to be critical for blocking the intermolecular binding between Rab27a and melanophilin.

Bioinformatics analysis of the epitope regions for norovirus capsid protein

BackgroundNorovirus is the major cause of nonbacterial epidemic gastroenteritis, being highly prevalent in both developing and developed countries. Despite of the available monoclonal antibodies (MAbs) for different sub-genogroups, a comprehensive epitope analysis based on various bioinformatics technology is highly desired for future potential antibody development in clinical diagonosis and treatment.MethodsA total of 18 full-length human norovirus capsid protein sequences were downloaded from GenBank. Protein modeling was performed with program Modeller 9.9. The modeled 3D structures of capsid protein of norovirus were submitted to the protein antigen spatial epitope prediction webserver (SEPPA) for predicting the possible spatial epitopes with the default threshold. The results were processed using the Biosoftware.ResultsCompared with GI, we found that the GII genogroup had four deletions and two special insertions in the VP1 region. The predicted conformational epitope regions mainly concentrated on N-terminal (1~96), Middle Part (298~305, 355~375) and C-terminal (560~570). We find two common epitope regions on sequences for GI and GII genogroup, and also found an exclusive epitope region for GII genogroup.ConclusionsThe predicted conformational epitope regions of norovirus VP1 mainly concentrated on N-terminal, Middle Part and C-terminal. We find two common epitope regions on sequences for GI and GII genogroup, and also found an exclusive epitope region for GII genogroup. The overlapping with experimental epitopes indicates the important role of latest computational technologies. With the fast development of computational immunology tools, the bioinformatics pipeline will be more and more critical to vaccine design.

Homology Modeling of Nitrogenase Iron Protein of Nitrogen Fixing Actinomycete Arthrobacter sp

Nitrogenase is an important enzyme associated with nitrogen fixation. Nitrogenase iron protein (Nif H) is one of the vital genes contributing to the nitrogen fixation. Arthrobacter sp. is one of the actinomycetes fixing atmospheric nitrogen. Nitrogenase iron protein of Arthrobacter sp. was retrieved from NCBI in FASTA format and a good template was selected by Basic Local Alignment Search tool and it was found to be Azotobacter vinelandii. The modeling of 3D structure of the protein was performed by Swiss model, GENO3D and ModWeb. The modeled 3D structure was evaluated by ProSA, VERIFY3D, PROCHECK, PROVE and ERRAT. The nitrogenase iron protein 3D structure modeled by ModWeb was found to be the best reliable model based on the above mentioned evaluation. The structural dynamics were performed by WEBnm@ and elNemo.3D structure of the nitrogenase iron protein was visualized by Rasmol.

Structure-Based Design and Screen of Novel Inhibitors for Class II 3-Hydroxy-3-methylglutaryl Coenzyme A Reductase from Streptococcus Pneumoniae

3-Hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) is a primary target in the current clinical treatment of hypercholesterolemia with specific inhibitors of "statin" family. Statins are excellent inhibitors of the class I (human) enzyme but relatively poor inhibitors of the class II enzyme, which are well-known as a potential target to discover drugs fighting against the invasive diseases originated from S. pneumoniae . However, no significantly effective inhibitors of class II HMGR have been reported so far. In the present study, the reasonable three-dimensional (3D) structure of class II HMGR from S. pneumoniae (SP-HMGR-II) was built by Swissmodel. On the basis of the modeling 3D structure in "close" flap domain form, several novel potential hit compounds out of SPECs database were picked out by using structure-based screening strategy. Especially the compounds 4, 3, and 11 exhibit highly inhibitory activities, with IC50 values of 11.5, 18.5, and 18.1 μM, respectively. Furthermore, the hit compounds were chosen as probe molecules, and their probable interactions with the corresponding individual residues have been examined by jointly using the molecular docking, site-directed mutagenesis, enzymatic assays, and fluorescence spectra, to provide an insight into a new special binding-model located between the HMG-CoA and NADPH pockets. The good agreement between theoretical and experimental results indicate that the modeling strategies and screening processes in the present study are very likely to be a promising way to search novel lead compounds with both structural diversity and high inhibitory activity against SP-HMGR-II in the future.

Amino acid features: a missing compartment of prediction of protein function

AbstractEnormous computational efforts have been carried out to predict structure and function of protein. However, nearly all of these efforts have been focused on prediction of function based on primary nucleic acid sequence or modeling 3D structure of protein from its nucleic acid sequence. In fact, it seems that amino acid attributes, which is an intermediate phase between DNA/RNA and advanced protein structure, has been missed.From 2010, we examined the possibility of precise prediction of structural protein function based on amino acid features by improving the following three aspects of amino acid research: (1) Increasing the number of computationally calculated amino acid features, (2) Testing different feature selection (attribute weighting) algorithms and selection of the most important amino acid attributes based on the overall conclusion of algorithms, (3) Examining different supervised and unsupervised data mining (machine learning) algorithms, and (4) Joining attribute weighting with different data mining algorithms. We applied the discovered procedure in different biological examples including: protein thermostability, halostability, prediction of function of heavy metal transporters, cancer diagnosis and prediction, and pursuing the EST-SSRs in amino acid level.In thermostability study, we successfully established an accurate expert system to predict the thermostability of any input sequence trough mining of its calculated amino acid features. Interestingly, performance of a clustering algorithm such as EMC can vary from 0.0% to 100%, depending upon which attribute weighting algorithm had summarized the attributes of the dataset prior to running the clustering algorithm.In another recent study on halostability, the results showed that amino acid composition can be used to efficiently discriminate halostable protein groups with up to 98% accuracy implying the possibility of precise prediction of halostability when an appropriate machine learning algorithm mines a large number of structural amino acid attributes of primary protein structure.Using our approach, simple amino acid features, without the need of advanced features of protein structure, could explain the difference between P1B-ATPases in hyperaccumulator and nonhyperaccumulator plants. More importantly, a precise model was built to discriminate P1B-ATPases in different organisms based on their structural amino acid features. In addition, for the first time, reliable models for prediction of the hyperaccumulating activity of unknown P1B-ATPase pumps were developed.We employed our method in monitoring and prediction of breast cancer. The results confirmed that amino acid composition can be used to discriminate between protein groups expressed in two forms of breast cancer: malignant and benign. This study was strong evidence that malignancy can be predicted out from amino acid, and malignant proteins can be distinguished based on the amino acid composition of their proteomes without further need for protein separation. An important outcome was the discovery of the role of dipeptides, in particular Ile-Ile, in cancer progression. In addition, Generalized Rule Induction (GRI) found association rules in the data showing the 100 most important rules classifying benign, malignant, and commonly expressed proteins expressed in breast cancers.In another investigation, we found that EST-SSRs in normal lung tissues are different than in unhealthy tissues, and tagged ESTs with SSRs cause remarkable differences in amino acid and protein expression patterns in cancerous tissue. This can be supposed as a glimpse of invention of a new sort of biomarkers based on frequency of amino acids.Up to now, phylogenic trees, drawn by nucleic acid or amino acid sequence alignments, have been employed as the base of evolutionary studies. However, this method does not take into account the structural and functional features of sequences during evolution. On the contrary, the presented classification here, based on the decision tree, anomaly detection model and feature weighting, provides an evolutionary separation of organisms based on their structural reasons of this diversity.Our findings have the potential to be efficiently used in the following area: filling the gap between laboratory engineering of proteins and computational biology, developing amino acid feature based-biomarkers, increasing the accuracy of prediction of 3D protein structure based on important amino acid features, and developing websites/software for prediction of the results of mutation. In addition, important discovered amino acid features can be employed as clues for discovering important DNA mutations and increasing prediction accuracy of 3D structure from DNA sequence. Furthermore, this study offers new for protein function, irrespective of similarity searches.

Studies on filarial GST as a target for antifilarial drug development—in silico and in vitro inhibition of filarial GST by substituted 1,4-naphthoquinones

Eleven 1,4-naphthoquinone analogues with different amino substitutions at position 3 of the quinone ring earlier reported for macrofilaricidal activity were selected and screened against purified cytosolic GST isolated from the bovine filarial worm Setaria digitata and IC(50) values were determined. Of the 11 compounds tested, 8 showed good inhibition against S. digitata GST. The IC(50) values of the most effective macrofilaricidal compounds-11 [2-(4-methylpiperazin-1-yl)naphthalene-1,4-dione] and 9 {2-[(1,3-dimethylbutyl)amino]naphthalene -1,4-dione}-were 0.872 and 0.994 mM, respectively. Compounds 9 and 11 were further studied for type of enzyme inhibition and found to exhibit competitive and uncompetitive inhibition kinetics, respectively, with respect to substrate GSH. All 11 compounds were in agreement with Lipinski's rule of five and passed through the FAFDrugs ADME/tox filter. Molecular docking was carried out using the modeled 3D structure of wbGST PDB ID:1SFM as receptor and substituted naphthoquinones as ligands using AutoDock 4.0. The binding energy of nine compounds varied from -9.15 to -6.58 Kcal mol(-1), whereas compounds 8 and 10 did not show any binding to the receptor. Among the compounds studied, compound 7 {2-[3-(diethylamino) propyl]aminonaphthalene-1,4-dione} showed maximum affinity towards wbGST as it exhibited the lowest binding energy, followed by compounds 11 and 9. However compound 7 was not macrofilaricidal while 11 and 9 exhibited macrofilaricidal activity. The results of in silico and in vitro studies with the synthesized 1,4 -naphthoquinone analogues on filarial GST and in vitro macrofilaricidal activity against adult bovine filarial worm S. digitata open up a promising biochemical target for antifilarial drug development.

SuperSweet--a resource on natural and artificial sweetening agents

A vast number of sweet tasting molecules are known, encompassing small compounds, carbohydrates, d-amino acids and large proteins. Carbohydrates play a particularly big role in human diet. The replacement of sugars in food with artificial sweeteners is common and is a general approach to prevent cavities, obesity and associated diseases such as diabetes and hyperlipidemia. Knowledge about the molecular basis of taste may reveal new strategies to overcome diet-induced diseases. In this context, the design of safe, low-calorie sweeteners is particularly important. Here, we provide a comprehensive collection of carbohydrates, artificial sweeteners and other sweet tasting agents like proteins and peptides. Additionally, structural information and properties such as number of calories, therapeutic annotations and a sweetness-index are stored in SuperSweet. Currently, the database consists of more than 8000 sweet molecules. Moreover, the database provides a modeled 3D structure of the sweet taste receptor and binding poses of the small sweet molecules. These binding poses provide hints for the design of new sweeteners. A user-friendly graphical interface allows similarity searching, visualization of docked sweeteners into the receptor etc. A sweetener classification tree and browsing features allow quick requests to be made to the database. The database is freely available at: http://bioinformatics.charite.de/sweet/.

The 3D-structural modeling of yeast D-amino acid oxidase

D-amino acid oxidase from the yeast Trigonopsis variabilis (TvDAAO) is used in the pharmaceutical industry and fine organic synthesis but for future practical applications new mutant forms of the enzyme with improved stability and catalytic properties are needed. Experiments on the crystallization of TvDAAO have been carried out for the last three decades without any success. For protein engineering of the enzyme using a rational design approach a model 3D-structure of TvDAAO was built using the homology modeling method. Cys108 and Cys298 residues were proposed for site-directed mutagenesis after analysis of the enzyme model structure.

Cracking pre-40S ribosomal subunit structure by systematic analyses of RNA–protein cross-linking

Understanding of eukaryotic ribosome synthesis has been slowed by a lack of structural data for the pre-ribosomal particles. We report rRNA-binding sites for six late-acting 40S ribosome synthesis factors, three of which cluster around the 3′ end of the 18S rRNA in model 3D structures. Enp1 and Ltv1 were previously implicated in ‘beak' structure formation during 40S maturation—and their binding sites indicate direct functions. The kinase Rio2, putative GTPase Tsr1 and dimethylase Dim1 bind sequences involved in tRNA interactions and mRNA decoding, indicating that their presence is incompatible with translation. The Dim1- and Tsr1-binding sites overlap with those of homologous Escherichia coli proteins, revealing conservation in assembly pathways. The primary binding sites for the 18S 3′-endonuclease Nob1 are distinct from its cleavage site and were unaltered by mutation of the catalytic PIN domain. Structure probing indicated that at steady state the cleavage site is likely unbound by Nob1 and flexible in the pre-rRNA. Nob1 binds before pre-rRNA cleavage, and we conclude that structural reorganization is needed to bring together the catalytic PIN domain and its target.

A Phylogenetic Approach for Assigning Function of Hypothetical Proteins in photorhabdus luminescens Subsp. laumondii TT01 Genome

Polyketides are larger family of structurally diverse natural products with a broad range of biological activity. The occurrence of polyketide synthase gene family/cluster in bacteria is capable to synthesis the polyketides. In this study, sequence and structural similarities of some hypothetical proteins of Photorhabdus luminescens subsp. laumondii TT01 analyzed to assign the functional relationship with polyketide synthases (PKSs) using bioinformatics tools. Many hypothetical proteins of this organism have shown homologies to PKS family on which a significant homolog found to be located at genomic region 2247626-2249476 bp in chromosome 1 carrying identical function. On searching motif and domain, it showed a strong similarity to ketoacyl (KA) synthase I and then to acyl carrier protein. The ketosynthaseacyltransferasedidomain module 5 (2HG4) of Saccharopolyspora erythraea found as a good ortholog and the best template for modeling 3D structure from the sequences of hypothetical proteins. ProFunc and Castp servers used to annotate the structure-function relationship of protein models. The structural aspects at primary and secondary levels also showed a close resemblance to KA synthase. Phylogenetic analysis of this sequence and protein model ensured its function would be
-KA synthase showing the functional reliability like ketosynthase, and it has evolutionary relationship with soil bacteria. There was a horizontal gene transfer event to acquire this domain in P.luminescence genome. Consequently, an abundance of PKS gene in the genome of entomopathogenic bacteria will obviously helpful to protect its host nematode from other pathological pervasiveness.

Characterization of biomimetc material, Mussel adhesive protein, using computer tools and servers.

In this paper six different mussel adhesion proteins (MAPs) retrieved from SwissProt database are analyzed using computer tools and servers. Primary analysis showed that all the MAPs are hydrophobic in nature due to the high content of non-polar amino acid residues. The sequence analysis showed the absence of sulphide bridges and non polar amino acid tryptophan in MAPs. Signal peptide is a common feature in MAPs. The aliphatic index computed by Expasy's Protparm infers that most of the MAPs are stable at a wide range of temperature. Secondary structure analysis showed that MAPs could results in a better interaction with water. The secondary structure analysis showed MAPs contain more βhelices and turns. The SOSUI server predicts one transmembarne region in Mytilus galprovincialis. The predicted transmembrane region was visualized and analyzed using helical wheel plots generated by EMBOSS PepWheel tool. The absence of disulphide bonds in MAPS was confirmed by SYC_REC tools and from 3 Dimensional structures created by Rasmol tool. The cystein position identified by Rasmol tool might be correct as the evaluated parameters (Rampage, CE and ProQ) are within the accepted limits for the model 3D structure. The signal peptide identified by SignalP server showed that they are secretary pathway in localization except in Mytilus edulus, where it is cytoplasmic. Based on the signal peptides reliability class the MAPs are classified into three groups. The phosphorylation sites of serine, threonoine and tyrosine are predicted by NetPhos server. Interestingly tyrosine is the most phosphorylated amino acid in the MAPs; hence it is inferred that MAPs can be purified by using antibodies.