MBU

Docking and pharmacophore screening tools were used to examine the binding of ligands in the active site of thymidine monophosphate kinase of Mycobacterium tuberculosis. Docking analysis of deoxythymidine monophosphate (dTMP) analogues suggests the role of hydrogen bonding and other weak interactions in enzyme selectivity. Water-mediated hydrogen-bond networks and a halogen-bond interaction seem to stabilize the molecular recognition. A pharmacophore model was developed using 20 dTMP analogues. The pharmacophoric features were complementary to the active site residues involved in the ligand recognition. On the basis of these studies, a composite screening model that combines the features from both the docking analysis and the pharmacophore model was developed. The composite model was validated by screening a database spiked with 47 known inhibitors. The model picked up 42 of these, giving an enrichment factor of 17. The validated model was used to successfully screen an in-house database of about 500 000 compounds. Subsequent screening with other filters gave 186 hit molecules.

Selective cyclooxygenase inhibitors have attracted much attention in recent times in the design of new non-steroidal anti-inflammatory drugs (NSAID). 3D-QSAR studies have been performed on a series of 1,5-diarylpyrazoles that act as selective cyclooxygenase-2 (COX-2) inhibitors, using three different methods: comparative molecular field analysis (CoMFA) with partial least squares (PLS) fit; molecular field analysis (MFA) and; receptor surface analysis (RSA) with genetic function algorithms (GFA). The analyses were carried out on 30 analogues of which 25 were used in the training set and the rest considered for the test set. These studies produced reasonably good predictive models with high cross-validated and conventional r 2 values in all the three cases.

Cyclic nucleotide phosphodiesterase IV (PDE IV) inhibitors find utility in asthma and Chronic Obstructive Pulmonary Disease (COPD) therapy. A series of 29 thieno[3,2-d]pyrimidines with affinity for PDE IV was subjected to three dimensional quantitative structure activity relationship (3D-QSAR) studies using comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA). Both CoMFA and CoMSIA provided statistically valid models with good correlative and predictive power. The incorporation of hydrophobic, hydrogen bond donor and hydrogen bond acceptor fields showed insignificant improvement in CoMSIA model. The 3D-QSAR models provide information for predicting the affinity of related compounds and designing more potent inhibitors. #

The 3D-QSAR studies of some indole derivatives as phosphodiesterase (PDE) type IV inhibitors was performed by Comparative Molecular Field Analysis (CoMFA) and Comparative Molecular Similarity Indices Analysis (CoMSIA) methods to determine the factors required for the activity of these compounds. The global minimum energy conformer of the template molecule, 3 the most active molecule of the series, was obtained by simulated annealing method and used to build the structures of the molecules in the dataset. The CoMFA model produced statistically significant results with cross-validated and conventional correlation coefficients of 0.494 and 0.986 respectively. The combination of steric, electrostatic and hydrophobic fields in CoMSIA gave the best results with cross-validated and conventional correlation coefficients of 0.541 and 0.967 respectively. The predictive ability of CoMFA and CoMSIA were determined using a test set of seven indole derivatives giving predictive correlation coefficients of 0.56 and 0.59 respectively indicating good predictive power. Further, the robustness of the models was verified by bootstrapping analysis. Based upon the information derived from CoMFA and CoMSIA, we have identified some key features that may be used to design new indole derivatives and predict their PDE IV affinities prior to synthesis.

deoxy-D-galaetose (GalNDns) is a useful fluorescent probe to study the interaction of non-fluorescent sugars with tile B4 lectin from Vicia villosa seeds (VVLB4). Binding of the lectin to GalNDns leads to a L2-fold increase in Dansyl fluorescence with a concomitant 10 nm blue shift in its emission maximum. The strong binding of GalNDns (/4, --7.33 × 10 ~ M -~ at 20°C) is due to a favourable entropic contribution to the association process. Among the other sugars studied, GalNAc,rl.O.Ser followed by Mer, GalNAc are the best ligands. 2-Deoxygalaetose, galacto~mine and galactose are 2013, 469 and 130 times weaker ligands, respectively, as compared Io GalNA¢, whereas GalNDns is about 2.44 times more potent than GalNAe. indicating that substitutions at the C-2 position of GalNAc have a considerable influence on the binding affinities. Equatorial orientation of the hydroxyl group at C-3 and axial orientation at C-4 as in galactose are important for the interaction with VVLB4. The C-6 hydroxyl group is not indispensable. The binding site of the leetin is directed exclusively towards monosaccharides alone. Interestingly enough, despite its preference Ibr Me,rGalNAc over MeflGalNAc, in oligosaccharides, the lectin prefers terminal ,8-1inked GaINAc as compared to the ~-linked one.

Three-dimensional quantitative structure-activity relationship (3D-QSAR) models were developed for 100 anilinoquinazolines, inhibiting epidermal growth factor receptor (EGFR) kinase. The studies included molecular field analysis (MFA) and receptor surface analysis (RSA). The cross-validated r 2 (r 2 cv ) values are 0.81 and 0.79 for MFA and RSA, respectively. The predictive ability of these models was validated by 28 test set molecules. The results of the best QSAR model were further compared with structure-based investigations using docking studies with the crystal structure of EGFR kinase domain. The results helped to understand the nature of substituents at the 6-and 7-positions, thereby providing new guidelines for the design of novel inhibitors. #

The hybrid pathway for heme biosynthesis in the malarial parasite proposes the involvement of parasite genome-coded enzymes of the pathway localized in different compartments such as apicoplast, mitochondria, and cytosol. However, knowledge on the functionality and localization of many of these enzymes is not available. In this study, we demonstrate that porphobilinogen deaminase encoded by the Plasmodium falciparum genome (PfPBGD) has several unique biochemical properties. Studies carried out with PfPBGD partially purified from parasite membrane fraction, as well as recombinant PfPBGD lacking N-terminal 64 amino acids expressed and purified from Escherichia coli cells (⌬PfPBGD), indicate that both the proteins are catalytically active. Surprisingly, PfPBGD catalyzes the conversion of porphobilinogen to uroporphyrinogen III (URO-GEN III), indicating that it also possesses uroporphyrinogen III synthase (UROS) activity, catalyzing the next step. This obviates the necessity to have a separate gene for UROS that has not been so far annotated in the parasite genome. Interestingly, ⌬PfP-BGD gives rise to UROGEN III even after heat treatment, although UROS from other sources is known to be heat-sensitive. Based on the analysis of active site residues, a ⌬PfPBGDL116K mutant enzyme was created and the specific activity of this recombinant mutant enzyme is 5-fold higher than ⌬PfPBGD. More interestingly, ⌬PfPBGDL116K catalyzes the formation of uroporphyrinogen I (UROGEN I) in addition to URO-GEN III, indicating that with increased PBGD activity the UROS activity of PBGD may perhaps become rate-limiting, thus leading to non-enzymatic cyclization of preuroporphyrinogen to URO-GEN I. PfPBGD is localized to the apicoplast and is catalytically very inefficient compared with the host red cell enzyme. parasite genomecoded PBGD lacking N-terminal 64 amino acids; PBS, phosphate-buffered saline; GFP, green fluorescent protein. Downloaded from FIGURE 5. Properties of native PBGD in the membrane fraction from P. falciparum. The membrane fraction was prepared from the parasite pellet, solubilized, and heat treated as described under "Experimental Procedures" and used for the assays. A and B, Western blot analysis of the membrane and cytosolic fractions from P. falciparum. M, marker; MF, membrane fraction; CY, cytosol fraction. C, time course. D, Lineweaver-Burk plot. E, HPLC profile of product. F, product spiked with URO III. G, product spiked with URO I.

Three dimensional quantitative structure-activity relationship (3D-QSAR) studies were carried out on deoxythymidine monophosphate (dTMP) derivatives inhibiting thymidine monophosphate kinase (TMPK) in Mycobacterium tuberculosis. Molecular field analysis (MFA) models with three different alignment techniques, namely, least squares, pharmacophore based and receptor based methods were developed. Receptor based MFA model showed better results when compared with least squares and pharmacophore based models. The results help us to understand the nature of substituents required for activity and thereby provide guidelines to design novel and potent inhibitors as antitubercular agents.

A comparative molecular field analysis (CoMFA) of phthalazine class of phosphodiesterase IV (PDE IV) inhibitors has been performed to correlate their chemical structures with their observed biological activity. A statistically valid model with good correlative and predictive power is reported. The leave one out cross-validation study gave cross-validation r 2 cv of value 0.507 at six optimum components and conventional r 2 of value 0.98. The predictive ability of the model was tested by predicting the seven molecules belonging to the test set giving predictive correlation coefficient of 0.59. This model is potentially helpful in the design of novel and more potent PDE IV inhibitors. #

In the models suggested recently for antigenic peptides binding in the ~, ~, groove of MHC class I molecules, the orientation of the peptide has been shown uniquely as: the N~C vector of the peptide being parallel to the N~C vector of the ~t~ helix of MHC, Here, v~e demonstrate ~hat the reverse orientation of the peptide is equally probable. This hypothesis is supported by molecular modelling calculations and computer graphic analyses on a routine class I MHC molecule H-SK ° and its complexes with a restricted peptide RYLENGKETLQ. Analysis of the complementary interactions between the peptid¢ residues and the amino acid side chains lining the MHC groove shows that the binding orientation of the peptide may be allele-specific and could depend on the sequence and structure of" the antigenic epitope.

A comparative molecular similarity indices analysis (CoMSIA) of a set of 29 imidazolyl and N-pyrrolyl heptenoates have been performed to find out the structural requirements for 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) inhibitory activity. The HMG like side chain, a common moiety of statins, was used to align the molecules. The results guide to design new chemical entities with high potency.

Riboswitches are structural cis-acting genetic regulatory elements in 59 UTRs of mRNAs, consisting of an aptamer domain that regulates the behavior of an expression platform in response to its recognition of, and binding to, specific ligands. While our understanding of the ligand-bound structure of the aptamer domain of the adenine riboswitches is based on crystal structure data and is well characterized, understanding of the structure and dynamics of the ligand-free aptamer is limited to indirect inferences from physicochemical probing experiments. Here we report the results of 15-nsec-long explicit-solvent molecular dynamics simulations of the add A-riboswitch crystal structure (1Y26), both in the adenine-bound (CLOSED) state and in the adenine-free (OPEN) state. Root-mean-square deviation, root-mean-square fluctuation, dynamic cross-correlation, and backbone torsion angle analyses are carried out on the two trajectories. These, along with solvent accessible surface area analysis of the two average structures, are benchmarked against available experimental data and are shown to constitute the basis for obtaining reliable insights into the molecular level details of the binding and switching mechanism. Our analysis reveals the interaction network responsible for, and conformational changes associated with, the communication between the binding pocket and the expression platform. It further highlights the significance of a, hitherto unreported, noncanonical W:H trans base pairing between A73 and A24, in the OPEN state, and also helps us to propose a possibly crucial role of U51 in the context of ligand binding and ligand discrimination.

Background: Cerebral malaria is a form of human malaria wherein Plasmodium falciparum-infected red blood cells adhere to the blood capillaries in the brain, potentially leading to coma and death. Interactions between parasite and host proteins are important in understanding the pathogenesis of this deadly form of malaria. It is, therefore, necessary to study available protein-protein interactions to identify lesser known interactions that could throw light on key events of cerebral malaria. Methods: Sequestration, haemostasis dysfunction, systemic inflammation and neuronal damage are key processes of cerebral malaria. Key events were identified from literature as being crucial to these processes. An integrated interactome was created using available experimental and predicted datasets as well as from literature. Interactions from this interactome were filtered based on Gene Ontology and tissue-specific annotations, and further analysed for relevance to the key events. Results: PfEMP1 presentation, platelet activation and astrocyte dysfunction were identified as the key events influencing the disease. 48896 host-parasite along with other host-parasite, host-host and parasite-parasite proteinprotein interactions obtained from a disease-specific corpus were combined to form an integrated interactome. Filtering of the interactome resulted in five host-parasite PPI, six parasite-parasite and two host-host PPI. The analysis of these interactions revealed the potential significance of apolipoproteins and temperature/Hsp expression on efficient PfEMP1 presentation; role of MSP-1 in platelet activation; effect of parasite proteins in TGF-β regulation and the role of albumin in astrocyte dysfunction. Conclusions: This work links key host-parasite, parasite-parasite and host-host protein-protein interactions to key processes of cerebral malaria and generates hypotheses for disease pathogenesis based on a filtered interaction dataset. These hypotheses provide novel and significant insights to cerebral malaria.

Comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) were performed on tetrahydrofuroyl-l-phenylalanine derivatives as VLA-4 antagonists. The best CoMFA and CoMSIA models that were generated using atom based alignment from a training set of twenty five tetrahydrofuroyl-l-phenylalanine derivatives, are sixcomponent models with good statistics; CoMFA: r 2 cv =0.366, r 2 =0.983, s=0.099, F=172.661 and PRESS=4.435; CoMSIA: r 2 cv =0.528, r 2 =0.995, s=0.054, F=577.87 and PRESS=3.563. Both of these 3-D-QSAR models were validated using a test set of eleven compounds, whose predicted pIC 50 values fall within one log unit of the actual pIC 50 . The contour diagrams obtained for the various CoMFA and CoMSIA field contributions can be mapped back onto structural features to explain the activity trends of the molecules analysed. Based on the spatial arrangement of the various field contributions, novel molecules with improved activity can be designed. #

-(2S)-pyrrolidine-2-methoxyl]phenylmethylene[thiazolidine-2,4-dione, malic acid salt]], a chemically distinct unsaturated thiazolidinedione, activates peroxisome proliferator-activated receptor ␥ (PPAR␥) submaximally in vitro with the binding affinity ϳ10 times less than that of rosiglitazone, a highly potent thiazolidinedione. PAT5A reduces plasma glucose level and improves insulin sensitivity in insulin resistant db/db mice, similar to that of rosiglitazone, while exerting a relatively weak adipogenic effect. In contrast to rosiglitazone, PAT5A inhibits cholesterol and fatty acid biosynthesis suggesting that PAT5A possesses a unique receptor-independent non-Article, publication date, and citation information can be found at

PDC-109, the major bovine seminal plasma protein, binds to sperm plasma membrane and modulates capacitation in the presence of heparin. In view of this, the PDC-109/heparin interaction has been investigated employing various biophysical approaches. Isothermal titration calorimetric studies yielded the association constant and changes in enthalpy and entropy for the interaction at 25°C (pH 7.4) as 1.92 ((0.2) Â 10 5 M À1 , 18.6 ((1.6) kcal M À1 , and 86.5 ((5.1) cal M À1 K À1 , respectively, whereas differential scanning calorimetric studies indicated that heparin binding results in a significant increase in the thermal stability of PDC-109. The affinity decreases with increase in pH and ionic strength, consistent with the involvement of electrostatic forces in this interaction. Circular dichroism spectroscopic studies indicated that PDC-109 retains its conformational features even up to 70À75°C in the presence of heparin, whereas the native protein unfolds at about 55°C. Atomic force microscopic studies demonstrated that large oligomeric structures are formed upon binding of PDC-109 to heparin, indicating an increase in the local density of the protein, which may be relevant to the ability of heparin to potentiate PDC-109 induced sperm capacitation.