Structure-based Drug Design Approach Research Articles

Molecular recognition of RNA: challenges for modelling interactions and plasticity

There is growing interest in molecular recognition processes of RNA because of RNA's widespread involvement in biological processes. Computational approaches are increasingly used for analysing and predicting binding to RNA, fuelled by encouraging progress in developing simulation, free energy and docking methods for nucleic acids. These developments take into account challenges regarding the energetics of RNA-ligand binding, RNA plasticity, and the presence of water molecules and ions in the binding interface. Accordingly, we will detail advances in force field and scoring function development for molecular dynamics (MD) simulations, free energy computations and docking calculations of nucleic acid complexes. Furthermore, we present methods that can detect moving parts within RNA structures based on graph-theoretical approaches or normal mode analysis (NMA). As an example of the successful use of these developments, we will discuss recent structure-based drug design approaches that focus on the bacterial ribosomal A-site RNA as a drug target.

In silico structure-based design of a novel class of potent and selective small peptide inhibitor of Mycobacterium tuberculosis Dihydrofolate reductase, a potential target for anti-TB drug discovery

The worldwide TB structural genomics initiative has identified several new drug targets for Mycobacterium tuberculosis (M. tb). Dihydrofolate reductase (DHFR) catalyzes the NADPH-dependent reduction of dihydrofolate to tetrahydrofolate that is essential for DNA synthesis. Inhibition of its activity leads to arrest of DNA synthesis and hence cell death. Thus, M. tb DHFR (mtDHFR) is an attractive novel drug target for developing anti-TB drugs. Structural comparison of mtDHFR and human DHFR (hDHFR) reveals key differences in the active sites. These differences can be exploited for the design of selective inhibitors for mtDHFR. Based on the recently determined high resolution crystal structure of mtDHFR complexed with known inhibitor methotrexate (MTX) and cofactor NADPH, a tri-peptide inhibitor has been identified using a structure-based drug design approach. Docking studies indicate that the designed tripeptide inhibitor has a high potency (K (d) = 1.78 nM) and is a selective (approximately 120 fold over hDHFR) inhibitor for mtDHFR. Hence, the tripeptide is a suitable lead compound for the development of novel anti-TB drugs.

Investigation into Adamantane-Based M2 Inhibitors with FB-QSAR

Because of their high resistance rate to the existing drugs, influenza A viruses have become a threat to human beings. It is known that the replication of influenza A viruses needs a pH-gated proton channel, the so-called M2 channel. Therefore, to develop effective drugs against influenza A, the most logic strategy is to inhibit the M2 channel. Recently, the atomic structure of the M2 channel was determined by NMR spectroscopy (Schnell, J.R. and Chou, J.J., Nature, 2008, 451, 591-595). The high-resolution NMR structure has provided a solid basis for structure-based drug design approaches. In this study, a benchmark dataset has been constructed that contains 34 newly-developed adamantane-based M2 inhibitors and covers considerable structural diversities and wide range of bioactivities. Based on these compounds, an in-depth analysis was performed with the newly developed fragment-based quantitative structure-activity relationship (FB-QSAR) algorithm. The results thus obtained provide useful insights for dealing with the drug-resistant problem and designing effective adamantane-based antiflu drugs.

Fragment‐based Identification of Hsp90 Inhibitors

Heat shock protein 90 (Hsp90) plays a key role in stress response and protection of the cell against the effects of mutation. Herein we report the identification of an Hsp90 inhibitor identified by fragment screening using a high-concentration biochemical assay, as well as its optimisation by in silico searching coupled with a structure-based drug design (SBDD) approach.

Structure-based design of molecular cancer therapeutics

Structure-based approaches now impact across the whole continuum of drug discovery, from new target selection through the identification of hits to the optimization of lead compounds. Optimal application of structure-based design involves close integration with other discovery technologies, including fragment-based and virtual screening. Here, we illustrate the use of structural information and of structure-based drug design approaches in the discovery of small-molecule inhibitors for cancer drug targets and provide an outlook on the exploitation of structural information in future cancer drug discovery. Examples include high profile protein kinase targets and structurally related PI3 kinases, histone deacetylases, poly(ADP-ribose)polymerase and the molecular chaperone HSP90. Structure-based design approaches have also been successfully applied to the protein-protein interaction targets p53-MDM2 and the Bcl-2 family.

CoMFA, LeapFrog and blind docking studies on sulfonanilide derivatives acting as selective aromatase expression regulators

Aromatase, the enzyme responsible for estrogen biosynthesis, is an attractive target in the treatment of hormone-dependent breast cancer. In this manuscript, the structure-based drug design approach of sulfonanilide analogues as potential selective aromatase expression regulators (SAERs) is described. Receptor-independent CoMFA (Comparative Molecular Field Analysis) maps were employed for generating a pseudocavity for LeapFrog calculation. A robust model, using 45 and 10 molecules in the training and test sets, respectively, was developed producing statistically significant results with cross-validated and conventional correlation coefficients of 0.656 and 0.956, respectively. This model was used to predict the activity of newly proposed molecules as SAERs candidates being two magnitude orders more potent than the previously reported compounds. Also in the present study, the computational blind docking method using eHiTS is tested on molecules study group and COX-2 enzyme. Future perspectives of the method in the screening of SAERs candidates with no COX-2 inhibitory activity are discussed.

Triclosan derivatives: towards potent inhibitors of drug-sensitive and drug-resistant Mycobacterium tuberculosis.

Triclosan has been previously shown to inhibit InhA, an essential enoyl acyl carrier protein reductase involved in mycolic acid biosynthesis, the inhibition of which leads to the lysis of Mycobacterium tuberculosis. Using a structure-based drug design approach, a series of 5-substituted triclosan derivatives was developed. Two groups of derivatives with alkyl and aryl substituents, respectively, were identified with dramatically enhanced potency against purified InhA. The most efficacious inhibitor displayed an IC(50) value of 21 nM, which was 50-fold more potent than triclosan. X-ray crystal structures of InhA in complex with four triclosan derivatives revealed the structural basis for the inhibitory activity. Six selected triclosan derivatives were tested against isoniazid-sensitive and resistant strains of M. tuberculosis. Among those, the best inhibitor had an MIC value of 4.7 microg mL(-1) (13 microM), which represents a tenfold improvement over the bacteriocidal activity of triclosan. A subset of these triclosan analogues was more potent than isoniazid against two isoniazid-resistant M. tuberculosis strains, demonstrating the significant potential for structure-based design in the development of next generation antitubercular drugs.

Novel Aldosterone Synthase Inhibitors with Extended Carbocyclic Skeleton by a Combined Ligand-Based and Structure-Based Drug Design Approach

Pharmacophore modeling of a series of aldosterone synthase (CYP11B2) inhibitors triggered the design of compounds 11 and 12 by extending a previously established naphthalene molecular scaffold (e.g., present in molecules 1 and 2) via introduction of a phenyl or benzyl residue in 3-position. These additional aromatic moieties have been hypothesized to fit into the newly identified hydrophobic pharmacophore feature HY3. Subsequent docking studies in our refined CYP11B2 protein model have been performed prior to synthesis to estimate the inhibitory properties of the proposed molecules. While phenyl-substituted compound 11 (IC50 > 500 nM) did not dock under the given pharmacophore constraint (i.e., the Fe(heme)-N(ligand) interaction), benzyl-substituted compound 12 (IC50 = 154 nM) was found to exploit a previously unexplored subpocket of the inhibitor binding site. By structural optimization based on the pharmacophore hypothesis, 25 novel compounds were synthesized, among them highly potent CYP11B2 inhibitors (e.g., 17, IC50 = 2.7 nM) with pronounced selectivity toward the most important steroidogenic and hepatic CYP enzymes.

Structural characterization of the involvement of SigC in the regulation of the gene expression of pathogenic Mycobacterium ulcerans

Mycobacterium ulcerans is the causative agent of the Buruli ulcers in humans. This disease is a devastating necrotic disease of the subcutaneous tissue. Currently there are very few drugs to prevent the Buruli ulcers. This human pathogen is carried by aquatic insects. The gene sequence of this pathogenic organism has recently been identified. In the present study an attempt has been made to analyze the structural characteristics of the SigC protein from Mycobacterium ulcerans. Sequence analyses of SigC reveal that it belongs to the family of sigma factors, which are involved in DNA binding at promoter sites, and play a role in transcription initiation. Homology modeling was employed to construct the three-dimensional structure of the protein and the model was analyzed. SigC protein has a distinct helix-turn-helix DNA binding motif. It is compared to other members of the DNA binding proteins. The modeled structure of SigC has been used to dock on the promoter DNA to predict the favourable binding interactions of the protein, which are responsible for the gene expressions in the organism. These interactions have been used to predict the plausible mechanism of the process. Since this is the first structural work regarding the interactions of SigC protein and its promoter DNA, this study may contribute in the realm of structure based drug design approaches to produce drugs against this harmful organism to control its proliferation and spreading of Buruli ulcer.

Design, synthesis and biological evaluation of bis(hydroxyphenyl) azoles as potent and selective non-steroidal inhibitors of 17β-hydroxysteroid dehydrogenase type 1 (17β-HSD1) for the treatment of estrogen-dependent diseases

The 17β-hydroxysteroid dehydrogenase type 1 (17β-HSD1) catalyses the reduction of the weakly active estrone (E1) into the most potent estrogen, 17β-estradiol (E2). E2 stimulates the growth of hormone-dependent diseases via activation of the estrogen receptors (ERs). 17β-HSD1 is often over-expressed in breast cancer cells. Thus, it is an attractive target for the treatment of mammary tumours. The combination of a ligand- and a structure-based drug design approach led to the identification of bis(hydroxyphenyl) azoles as potential inhibitors of 17β-HSD1. Different azoles and hydroxy substitution patterns were investigated. The compounds were evaluated for activity and selectivity with regard to 17β-HSD2, ERα and ERβ. The most potent compound is 3-[5-(4-hydroxyphenyl)-1,3-oxazol-2-yl]phenol ( 18, IC 50 = 0.31 μM), showing very good selectivity, high cell permeability and medium CaCo-2 permeability.

Challenges in Modern Drug Discovery: A Case Study of Boceprevir, an HCV Protease Inhibitor for the Treatment of Hepatitis C Virus Infection

More than 170 million people worldwide are affected by the hepatitis C virus (HCV). The disease has been described as a "silent epidemic" and "a serious global health crisis". HCV infection is a leading cause of chronic liver disease such as cirrhosis, carcinoma, or liver failure. The current pegylated interferon and ribavirin combination therapy is effective in only 50% of patients. Its moderate efficacy and apparent side effects underscore the need for safer and more effective treatments. The nonstructural NS3 protease of the virus plays a vital role in the replication of the HCV virus. The development of small molecule inhibitors of NS3 protease as antiviral agents has been intensively pursued as a viable strategy to eradicate HCV infection. However, it is a daunting task. The protease has a shallow and solvent-exposed substrate binding region, and the inhibitor binding energy is mainly derived from weak lipophilic and electrostatic interactions. Moreover, lack of a robust in vitro cell culture system and the absence of a convenient small animal model have hampered the assessment of both in vitro and in vivo efficacy of any antiviral compounds. Despite the tremendous challenges, with access to a recently developed cell-based replicon system, major progress has been made toward a more effective small molecule HCV drug. In our HCV program, facing no leads from our screening effort, a structure-based drug design approach was carried out. An alpha-ketoamide-type electrofile was designed to trap the serine hydroxyl of the protease. Early ketoamide inhibitors mimicked the structures of the peptide substrates. With the aid of X-ray structures, we successfully truncated the undecapeptide lead that had a molecular weight of 1265 Da stepwise to a tripeptide with a molecular weight of 500 Da. In an attempt to depeptidize the inhibitors, various strategies such as hydrazine urea replacement of amide bonds and P2 to P4 and P1 to P3 macrocyclizations were examined. Further optimization of the tripeptide inhibitors led to the identification of the best moieties for each site: primary ketoamide at P', cyclobutylalanine at P1, gem-dimethylcyclopropylproline at P2, tert-leucine at P3, and tert-butyl urea as capping agent. The combination of these led to the discovery of compound 8 (SCH 503034, boceprevir), our clinical candidate. It is a potent inhibitor in both enzyme assay (Ki* = 14 nM) and cell-based replicon assay (EC 90 = 0.35 microM). It is highly selective (2200x) against human neutrophil elastase (HNE). Boceprevir is well tolerated in humans and demonstrated antiviral activity in phase I clinical trials. It is currently in phase II trials. This Account details the complexity and challenges encountered in the drug discovery process.

Activation of JAZ by Small Molecules in Human Leukemia Cells.

By screening a murine interleukin-3 (IL-3)-dependent myeloid cell cDNA library, we previously identified JAZ (Just Another Zinc Finger Protein), a novel zinc finger protein. JAZ belongs to a new class of evolutionarily conserved C2H2-type ZFPs that feature an unusually long linker sequence and preferentially bind to double-stranded RNA. JAZ localizes in the nucleus and its human gene is localized to the chromosome 5q35-qter, a specific chromosomal region at which deletions and translocations occur in leukemia and lymphoma. We have recently discovered JAZ as a novel direct, positive regulator of p53 transcriptional activity. The mechanism involves direct binding to p53's C-terminal (negative) regulatory domain to activate “latent” p53 in response to non-genotoxic stress signals (such as interleukin-3 growth factor withdrawal). Thus, we have explored JAZ as a potentially novel molecular target in human leukemia by identifying small molecules to activate JAZ-mediated leukemic cell killing. A structure-based drug design approach was employed to screen small molecules that potentially bind and activate JAZ. Cytotoxicity assays were then carried out with a set of candidate compounds in a dose- and time-dependent manner in various human leukemia cell lines including REH, HL-60, U937, HEL and K562 cells. Several JAZ “activating” compounds were selected, which display differential abilities to induce leukemic cell killing (IC50 = < 1 to 100 μM). Interestingly, biochemical analysis shows that the J-compound(s) can mediate cell death in p53 expressing leukemia cells in association with upregulation of expression of JAZ, p53 and the proapoptotic p53 target gene BAX, indicating activation of p53. Furthermore, the J-compound(s) synergizes induction of leukemic cell death when combined with other known p53 activating agents including cisplatin and nutlin-3, a recently developed small molecule MDM2 antagonist that disrupts the p53-MDM2 interaction. Therefore, the J-compound(s) may mediate leukemic cell death in a mechanism involving activation of “latent”, wild-type p53 by targeting JAZ. However, the J-compound-mediated cell death was also observed in some p53-deficient leukemia cell lines. This suggests that depending on the cell type the J-compounds may also act by a p53-independent mechanism. Since JAZ was recently reported to be a cargo protein for exportin-5, the nuclear export receptor for pre-microRNAs, this dsRNA-binding ZFP may also have an unknown p53-independent function via which the J-compound(s) may possibly act. While these possible mechanisms remain to be further investigated, the J-compound(s) points the way to develop a potentially novel therapeutic strategy targeting JAZ to treat human leukemia.

DCC-2036: A Novel Switch Pocket Inhibitor of ABL Tyrosine Kinase with Therapeutic Efficacy Against BCR-ABL T315I In Vitro and in a CML Mouse Model.

Therapy with ATP-competitive ABL kinase inhibitors, including imatinib (Gleevec), dasatinib (Sprycel), and nilotinib (Tasigna), has revolutionized the treatment of chronic myeloid leukemia (CML), but a substantial proportion of patients develop resistance to these agents. A major mechanism of acquired resistance is mutations in the ABL kinase domain that render BCR-ABL insensitive to the drug; in particular, mutation of the ABL kinase “gatekeeper” Thr315 residue to Ile (T315I) confers pan-resistance to all ATP-competitive ABL inhibitors. To address this need, we developed and characterized a novel chemical class of compounds that bind to five structural pockets involved in the endogenous “switch” mechanism used by the ABL kinase to conformationally control its activity state. Using a structure-based drug design approach, diversity in these “switch pockets” between kinases can be exploited to develop inhibitors with high potency and specificity. The resulting ABL inhibitors have several attractive features, as they:1.act through a non-ATP-competitive mechanism;2.avoid steric clash with Ile315;3.inhibit purified ABL that is either unphosphorylated (switch-off) or phosphorylated (switch-on) at Tyr393 with IC50 of 0.8–4.0 nM;4.co-crystallize with phospho-Y393-ABL WT and T315I; and5.have very prolonged residency time on the kinase (off-rate ∼400 min vs. 3 min for imatinib).A lead compound, DCC-2036, is highly selective for ABL, FLT3, and SRC family kinases, and has favorable pharmacokinetic and toxicity profiles in animals. In vitro, DCC-2036 inhibited proliferation and induced apoptosis of Ba/F3 cells expressing BCR-ABL WT or several common imatinib-resistant mutants (Y253F, T315I, M351T) with IC50 from 5 to 25 nM, without appreciable inhibition of parental Ba/F3 cells in IL-3 (IC50 >1000 nM). In Balb/c mice injected with Ba/F3-BCR-ABL T315I cells, pharmacodynamic studies indicated >8 hour inhibition of phospho-Stat5 and phospho-ABL in the leukemic cells following a single oral 100 mg/kg dose of DCC-2036, whereas daily dosing of DCC-2036 significantly prolonged survival. In the mouse retroviral bone marrow transduction/transplantation model of CML, DCC-2036 at 100 mg/kg/d also reduced peripheral blood leukocyte counts and significantly prolonged survival vs. vehicle-treated control mice. These results demonstrate that ABL switch pocket inhibitors are a promising new pharmacologic weapon for the treatment of de novo and drug-resistant CML, including BCR-ABL T315I.

DrugScoreRNAKnowledge-Based Scoring Function To Predict RNA−Ligand Interactions

There is growing interest in RNA as a drug target due to its widespread involvement in biological processes. To exploit the power of structure-based drug-design approaches, novel scoring and docking tools need to be developed that can efficiently and reliably predict binding modes and binding affinities of RNA ligands. We report for the first time the development of a knowledge-based scoring function to predict RNA-ligand interactions (DrugScoreRNA). Based on the formalism of the DrugScore approach, distance-dependent pair potentials are derived from 670 crystallographically determined nucleic acid-ligand and -protein complexes. These potentials display quantitative differences compared to those of DrugScore (derived from protein-ligand complexes) and DrugScoreCSD (derived from small-molecule crystal data). When used as an objective function for docking 31 RNA-ligand complexes, DrugScoreRNA generates "good" binding geometries (rmsd (root mean-square deviation) < 2 A) in 42% of all cases on the first scoring rank. This is an improvement of 44% to 120% when compared to DrugScore, DrugScoreCSD, and an RNA-adapted AutoDock scoring function. Encouragingly, good docking results are also obtained for a subset of 20 NMR structures not contained in the knowledge-base to derive the potentials. This clearly demonstrates the robustness of the potentials. Binding free energy landscapes generated by DrugScoreRNA show a pronounced funnel shape in almost 3/4 of all cases, indicating the reduced steepness of the knowledge-based potentials. Docking with DrugScoreRNA can thus be expected to converge fast to the global minimum. Finally, binding affinities were predicted for 15 RNA-ligand complexes with DrugScoreRNA. A fair correlation between experimental and computed values is found (RS = 0.61), which suffices to distinguish weak from strong binders, as is required in virtual screening applications. DrugScoreRNA again shows superior predictive power when compared to DrugScore, DrugScoreCSD, and an RNA-adapted AutoDock scoring function.

Design, Synthesis, and Performance of NTA-modified Lipids as Templates for Histidine-tagged Protein Crystallization

Abstract Current methods of drug discovery and development typically involve either structure-based drug design approaches or high-throughput screening methods for lead identification. The structure-based discovery process relies heavily upon the labor-intensive determination of three-dimensional protein structures using X-ray diffraction (XRD) and NMR techniques. Although tens of thousands of soluble protein structures have been resolved at the atomic level using these methods, there are many drug targets, particularly integral membrane proteins, whose structures have not been elucidated. In the case of XRD, this limitation often arises from the lack of reliable and predictable methods for protein crystallization. The extensive number of proteins crystallized by the interfacial templating approach suggests that this method may offer great promise for structural biology applications, particularly for proteins that do not readily form X-ray quality crystals from bulk solution. This review highlights the various approaches that have been used for templating of protein crystallization at gas–liquid and solid–liquid interfaces. Special emphasis is placed on the synthesis of NTA- and IDA-conjugated lipids, their use in templating and concentrating proteins at lipid monolayer interfaces, and protein structure elucidations that have been facilitated by these metal-chelating lipids. Current approaches to control the nucleation and growth of protein crystals involving microfluidics, dip pen nanolithography, and non-covalent symmetry-based templating approaches are also discussed. Taken together, the progress in this field suggests that the interfacial templating approach to crystallization can develop into a powerful tool for high-throughput structural analysis by providing a universal, readily controllable method for crystallizing proteins.

Megsin Gene: Its Genomic Analysis, Pathobiological Functions, and Therapeutic Perspectives

It is critical to uncover genes specifically expressed in individual cell types for further understanding of cell biology and pathology. In order to elucidate pathogenesis of renal disease, we performed functional quantitative analysis of the genome in human kidney cells and compared the expression levels of a variety of kidney transcripts with those in other non-kidney cells. As a result, we identified a novel human gene, megsin, which is a new serine protease inhibitor (serpin) predominantly expressed in the kidney. Megsin is up-regulated in kidney disease. Genomic analysis revealed an association of the polymorphisms of megsin gene with susceptibility and/or progression of kidney disease. Its overexpression in rodents has led to the recognition of two different kidney abnormalities. The first disorder is linked to megsin biological effect itself and the other to its conformational abnormality recently called the serpinopathy. In the latter model, the cellular and tissue damage is induced by the endoplasmic reticulum (ER) stress due to conformational disorder resulting from megsin tertiary structure. In both types, the inhibition of megsin's activity or abnormal conformational change should open new therapeutic perspectives. The desire to prevent these abnormalities with the hope to offer new therapeutic strategies has stimulated the development of megsin inhibitors by a structure based drug design approach relying on a precisely known three dimensional megsin structure.

Ligand-Based Homology Modeling as Attractive Tool to Inspect GPCR Structural Plasticity

G protein-coupled receptors (GPCRs) represent the largest family known of signal-transducing molecules. They convey signals for light and many extracellular regulatory molecules. GPCRs have been found to be dysfunctional/dysregulated in a growing number of human diseases and they have been estimated to be the targets of more than 40% of the drugs used in clinical medicine today. The crystal structure of rhodopsin provides the first three-dimensional GPCR information, which now supports homology modeling studies and structure-based drug design approaches. Here, we review our recent work on adenosine receptors, a family of GPCRs and, in particular, on A(3) adenosine receptor subtype antagonists. We will focus on an alternative approach to computationally explore the multi-conformational space of the antagonist-like state of the human A(3) receptor. We define ligand-based homology modeling as new approach to simulate the reorganization of the receptor induced by the ligand binding. The success of this approach is due to the synergic interaction between theory and experiment.

Targeting Cancer: The Challenges and Successes of Structure-Based Drug Design Against the Human Purinome

Purine-binding proteins are of critical importance to all living organisms. Approximately 13% of the human genome is devoted to coding for purine-binding proteins. Given their importance, purine-binding proteins are attractive targets for chemotherapeutic intervention against a variety of disease states, particularly cancer. Modern computational and biophysical techniques, combined together in a structure-based drug design approach, aid immensely in the discovery of inhibitors of these targets. This review covers the process of modern structure-based drug design and gives examples of its use in discovery and development of drugs that target purine-binding proteins. The targets reviewed are human purine nucleoside phosphorylase, human epidermal growth factor receptor kinase, and human kinesin spindle protein.

Elucidation of sulfadoxine resistance with structural models of the bifunctional Plasmodium falciparum dihydropterin pyrophosphokinase–dihydropteroate synthase

Resistance of the most virulent human malaria parasite, Plasmodium falciparum, to antifolates is spreading with increasing speed, especially in Africa. Antifolate resistance is mainly caused by point mutations in the P. falciparum dihydropteroate synthase (DHPS) and dihydrofolate reductase (DHFR) target proteins. Homology models of the bifunctional P. falciparum dihydropterin pyrophosphokinase–dihydropteroate synthase (PPPK–DHPS) enzyme as well as the separate domains complete with bound substrates were constructed using the crystal structures of Saccharomyces cerevisiae (PPPK–DHPS), Mycobacterium tuberculosis (DHPS), Bacillus anthracis (DHPS), and Escherichia coli (PPPK) as templates. The resulting structures were subsequently solvated and refined using molecular dynamics. The active site residues of DHPS are highly conserved in S. cerevisiae, M. tuberculosis, E. coli, S. aureus, and B. anthracis, an attribute also shared by P. falciparum DHPS. Sulfadoxine was superimposed into the equivalent position of the p-aminobenzoic acid substrate and its binding parameters were refined using minimization and molecular dynamics. Sulfadoxine appears to interact mainly with P. falciparum DHPS mainly through hydrophobic interactions. Rational explanations are provided by the model for the sulfadoxine resistance-causing effects of four of the five known mutations in P. falciparum DHPS. A possible structure for the bifunctional PPPK–DHPS was derived from the structure from the S. cerevisiae bifunctional enzyme. The active site residues of P. falciparum PPPK are also conserved when compared to S. cerevisiae, Haemophilus influenzae, and E. coli. The informative nature of these models opens up avenues for structure-based drug design approaches toward the development of alternative and more effective inhibitors of P. falciparum PPPK–DHPS.

Structure-based drug design meets the ribosome

The high-resolution structures of the bacterial ribosomal subunits and those of their complexes with antibiotics have advanced significantly our understanding of small-molecule interactions with RNA. The wealth of RNA structural data generated by these structures has allowed computational chemists to employ a drug discovery paradigm focused on RNA-based targets. The structures also show how target-based resistance affects antibiotics acting at the level of the ribosome. Not only are the sites pinpointed where different classes of antibiotics inhibit protein synthesis, but their orientations, relative dispositions, and unique mechanisms of action are also revealed at the atomic level. Both the 30S and the 50S ribosomal subunits have been shown to be “targets of targets”, offering several adjacent, functionally relevant binding pockets for antibiotics. It is the detailed knowledge of these validated locations, or ribofunctional loci, plus the mapping of the resistance hot-spots that allow the rational design of next-generation antibacterials. When the structural information is combined with a data-driven computational toolkit able to describe and predict molecular properties appropriate for bacterial cell penetration and drug-likeness, a structure-based drug design approach for novel antibacterials shows great promise.