The structure of PknB in complex with mitoxantrone, an ATP-competitive inhibitor, suggests a mode of protein kinase regulation in mycobacteria

Unité de Biochimie Structurale and CNRS URA2185, Institut Pasteur, 25 rue du Docteur Roux, 75724 Paris, France.
FEBS Letters (Impact Factor: 3.17). 06/2006; 580(13):3018-22. DOI: 10.1016/j.febslet.2006.04.046
Source: PubMed


Mycobacterium tuberculosis PknB is an essential receptor-like protein kinase involved in cell growth control. Here, we demonstrate that mitoxantrone, an anthraquinone derivative used in cancer therapy, is a PknB inhibitor capable of preventing mycobacterial growth. The structure of the complex reveals that mitoxantrone partially occupies the adenine-binding pocket in PknB, providing a framework for the design of compounds with potential therapeutic applications. PknB crystallizes as a 'back-to-back' homodimer identical to those observed in other structures of PknB in complex with ATP analogs. This organization resembles that of the RNA-dependent protein kinase PKR, suggesting a mechanism for kinase activation in mycobacteria.

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    • "MATERIALS AND METHODS Protein structure retrieval: The 3D structure of cytosolic domain of the PKnB Ser/Thr Kinase receptor protein from Mycobacterium tuberculosis was retrieved from RCSB Protein Data Bank (PDB). The protein model with PDB ID: 2FUM [1] was chosen for active site predictions and further docking studies. "
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    ABSTRACT: Ser/Thr Kinase is one of the four M. tuberculosis kinases that are conserved in the downsized genome of Mycobacterium leprae and are therefore presumed to play an important role in the processes that regulate the complex life cycle of mycobacteria. It is known that there are two main superfamilies of protein kinases, one including STPKs1 and PTKs and that of His kinases. For a long time, the former were only found in eukaryotes, and the latter were only found in prokaryotes. In this paradigm, proteins from each superfamily were supposed to play analogous roles in the essentially different organization of signal transduction in both phyla. In this study we report the binding mode of Ser/Thr kinase with derivatives of Betulin ((lup-20(29)-ene-3β,28-diol) on the basis of structural similarity, substructure, isomers & conformers. Molecular docking approach using Lamarckian Genetic Algorithm was carried out to find out the potent inhibitors for Ser/Thr Kinase on the basis of calculated ligand-protein pairwise interaction energies. Study was carried out on 3000 molecules which were virtually screened from different databases on the basis of the structural similarity of Betulin. The grid maps representing the protein were calculated using auto grid and grid size was set to 60*60*60 points with grid spacing of 0.375 Ǻ. Docking was carried out with standard docking protocol on the basis of a population size of 150 randomly placed individuals; a maximum number of 2.5 *107 energy evaluations, a mutation rate of 0.02, a crossover rate of 0.80 and an elitism value of 1. Fifteen independent docking runs were carried out for each ligand and results were clustered according to the 1.0 Ǻ rmsd criteria. The docking result of the study of 3000 molecules demonstrated that the binding energies were in the range of -12.72 kcal/mol to -1.71 kcal/mol, with the minimum binding energy of –12.72 kcal/mol. 6 molecules showing hydrogen bonds with the active site residue VAL 95. Further in-vitro and in-vivo study is required on these molecules as the binding mode provided hints for the future design of new derivatives with higher potency and specificity.
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    • "In this work we have used ligand and structure based approaches to screen large set of inhibitors. Previously many high affinity inhibitors have been reported for PknB [11-15]: we used 62 inhibitors listed in Additional file 1 for our work. "
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    ABSTRACT: Background: Mycobacterium tuberculosis encodes 11 putative serine-threonine proteins Kinases (STPK) which regulates transcription, cell development and interaction with the host cells. From the 11 STPKs three kinases namely PknA, PknB and PknG have been related to the mycobacterial growth. From previous studies it has been observed that PknB is essential for mycobacterial growth and expressed during log phase of the growth and phosphorylates substrates involved in peptidoglycan biosynthesis. In recent years many high affinity inhibitors are reported for PknB. Previously implementation of data fusion has shown effective enrichment of active compounds in both structure and ligand based approaches .In this study we have used three types of data fusion ranking algorithms on the PknB dataset namely, sum rank, sum score and reciprocal rank. We have identified reciprocal rank algorithm is capable enough to select compounds earlier in a virtual screening process. We have also screened the Asinex database with reciprocal rank algorithm to identify possible inhibitors for PknB. Results: In our work we have used both structure-based and ligand-based approaches for virtual screening, and have combined their results using a variety of data fusion methods. We found that data fusion increases the chance of actives being ranked highly. Specifically, we found that the ranking of Pharmacophore search, ROCS and Glide XP fused with a reciprocal ranking algorithm not only outperforms structure and ligand based approaches but also capable of ranking actives better than the other two data fusion methods using the BEDROC, robust initial enhancement (RIE) and AUC metrics. These fused results were used to identify 45 candidate compounds for further experimental validation. Conclusion: We show that very different structure and ligand based methods for predicting drug-target interactions can be combined effectively using data fusion, outperforming any single method in ranking of actives. Such fused results show promise for a coherent selection of candidates for biological screening.
    Journal of Cheminformatics 01/2013; 5(1):2. DOI:10.1186/1758-2946-5-2 · 4.55 Impact Factor
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    • "A total of 54 000 compounds (diverse compound and kinase-focused collection) were tested measuring the in vitro phosphorylation of GarA (Rv1827) by PknB [81,82]. This study has permitted the identification of a new class of ATP competitive inhibitors with IC50s in the nM range [27]. However, improved inhibitors are still awaited, since their MICs values against Mtb growth within macrophages were found to be only in the micromolar range, likely due to limited cell wall permeability [27]. "
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    ABSTRACT: Recent genetic, biochemical and structural studies have established that eukaryotic-like Ser/Thr protein-kinases are critical mediators of developmental changes and host pathogen interactions in bacteria. Although with lower abundance compared to their homologues from eukaryotes, Ser/Thr protein-kinases are widespread in gram-positive bacteria. These data underline a key role of reversible Ser/Thr phosphorylation in bacterial physiology and virulence. Numerous studies have revealed how phosphorylation/dephosphorylation of Ser/Thr protein-kinases governs cell division and cell wall biosynthesis and that Ser/Thr protein kinases are responsible for distinct phenotypes, dependent on different environmental signals. In this review we discuss the current understandings of Ser/Thr protein-kinases functional processes based on structural data.
    Current Protein and Peptide Science 12/2012; 13(8). DOI:10.2174/138920312804871201 · 3.15 Impact Factor
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