Dataset Information

Structure of MurF from Streptococcus pneumoniae co-crystallized with a small molecule inhibitor exhibits interdomain closure.

ABSTRACT: In a broad genomics analysis to find novel protein targets for antibiotic discovery, MurF was identified as an essential gene product for Streptococcus pneumonia that catalyzes a critical reaction in the biosynthesis of the peptidoglycan in the formation of the cell wall. Lacking close relatives in mammalian biology, MurF presents attractive characteristics as a potential drug target. Initial screening of the Abbott small-molecule compound collection identified several compounds for further validation as pharmaceutical leads. Here we report the integrated efforts of NMR and X-ray crystallography, which reveal the multidomain structure of a MurF-inhibitor complex in a compact conformation that differs dramatically from related structures. The lead molecule is bound in the substrate-binding region and induces domain closure, suggestive of the domain arrangement for the as yet unobserved transition state conformation for MurF enzymes. The results form a basis for directed optimization of the compound lead by structure-based design to explore the suitability of MurF as a pharmaceutical target.

SUBMITTER: Longenecker KL

PROVIDER: S-EPMC2253247 | biostudies-literature | 2005 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Structure of MurF from Streptococcus pneumoniae co-crystallized with a small molecule inhibitor exhibits interdomain closure.

Longenecker Kenton L KL Stamper Geoffrey F GF Hajduk Philip J PJ Fry Elizabeth H EH Jakob Clarissa G CG Harlan John E JE Edalji Rohinton R Bartley Diane M DM Walter Karl A KA Solomon Larry R LR Holzman Thomas F TF Gu Yu Gui YG Lerner Claude G CG Beutel Bruce A BA Stoll Vincent S VS

Protein science : a publication of the Protein Society 20051201 12

In a broad genomics analysis to find novel protein targets for antibiotic discovery, MurF was identified as an essential gene product for Streptococcus pneumonia that catalyzes a critical reaction in the biosynthesis of the peptidoglycan in the formation of the cell wall. Lacking close relatives in mammalian biology, MurF presents attractive characteristics as a potential drug target. Initial screening of the Abbott small-molecule compound collection identified several compounds for further vali ...[more]

PMID: 16322581

Similar Datasets

Project description:Streptococcus pneumoniae persist in the human nasopharynx within organized biofilms. However, expansion to other tissues may cause severe infections such as pneumonia, otitis media, bacteremia, and meningitis, especially in children and the elderly. Bacteria within biofilms possess increased tolerance to antibiotics and are able to resist host defense systems. Bacteria within biofilms exhibit different physiology, metabolism, and gene expression profiles than planktonic cells. These differences underscore the need to identify alternative therapeutic targets and novel antimicrobial compounds that are effective against pneumococcal biofilms. In bacteria, DNA adenine methyltransferase (Dam) alters pathogenic gene expression and catalyzes the methylation of adenine in the DNA duplex and of macromolecules during the activated methyl cycle (AMC). In pneumococci, AMC is involved in the biosynthesis of quorum sensing molecules that regulate competence and biofilm formation. In this study, we examine the effect of a small molecule Dam inhibitor, pyrimidinedione, on Streptococcus pneumoniae biofilm formation and evaluate the changes in global gene expression within biofilms via microarray analysis. The effects of pyrimidinedione on in vitro biofilms were studied using a static microtiter plate assay, and the architecture of the biofilms was viewed using confocal and scanning electron microscopy. The cytotoxicity of pyrimidinedione was tested on a human middle ear epithelium cell line by CCK-8. In situ oligonucleotide microarray was used to compare the global gene expression of Streptococcus pneumoniae D39 within biofilms grown in the presence and absence of pyrimidinedione. Real-time RT-PCR was used to study gene expression. Pyrimidinedione inhibits pneumococcal biofilm growth in vitro in a concentration-dependent manner, but it does not inhibit planktonic cell growth. Confocal microscopy analysis revealed the absence of organized biofilms, where cell-clumps were scattered and attached to the bottom of the plate when cells were grown in the presence of pyrimidinedione. Scanning electron microscopy analysis demonstrated the absence of an extracellular polysaccharide matrix in pyrimidinedione-grown biofilms compared to control-biofilms. Pyrimidinedione also significantly inhibited MRSA, MSSA, and Staphylococcus epidermidis biofilm growth in vitro. Furthermore, pyrimidinedione does not exhibit eukaryotic cell toxicity. In a microarray analysis, 56 genes were significantly up-regulated and 204 genes were significantly down-regulated. Genes involved in galactose metabolism were exclusively up-regulated in pyrimidinedione-grown biofilms. Genes related to DNA replication, cell division and the cell cycle, pathogenesis, phosphate-specific transport, signal transduction, fatty acid biosynthesis, protein folding, homeostasis, competence, and biofilm formation were down regulated in pyrimidinedione-grown biofilms. This study demonstrated that the small molecule Dam inhibitor, pyrimidinedione, inhibits pneumococcal biofilm growth in vitro at concentrations that do not inhibit planktonic cell growth and down regulates important metabolic-, virulence-, competence-, and biofilm-related genes. The identification of a small molecule (pyrimidinedione) with S. pneumoniae biofilm-inhibiting capabilities has potential for the development of new compounds that prevent biofilm formation.

Project description:Streptococcus pneumoniae is a Gram positive bacterium that causes severe invasive infection such as pneumonia, septicemia, meningitis and otitis media especially in children, the elderly and immune-compromised patients. Pneumococcal colonization and disease is often associated with biofilm formation. Bacteria in biofilms exhibit elevated resistance both to antibiotics and to host defense systems, which often results in persistent and difficult-to-treat infections. Therefore, the ongoing treat to human health posed by pneumococcal biofilms has prompted extensive research aimed to identify alternative targets and new antimicrobial agents that are effective against bacteria biofilms. The effective anti-biofilm strategies should include inhibition of microbial adhesion to the surface and of colonization, interference with the signal molecules modulating biofilm development and the disaggregation of the biofilm matrix. In this study, we examine the effect of DAM inhibitor small molecule pyrimidine-diones on streptococcus pneumoniae D-39 strain growth (planktonic and biofilm) and evaluate the changes in global gene expression using c-DNA microarray. The microarray analysis was performed on total RNA extracted from biofilms grown in 24-well microtiter plate with 7µm/ml pyrimidine-diones small molecule and control biofilms (biofilms grown without pyrimidine-diones small molecule). To validate the results of microarray, real-time RT-PCR was performed on 12 differentially expressed genes from six different functional groups. cDNA-microarray analysis detected a total of 259 genes that were significantly differentially expressed in biofilm growth with pyrimidine-diones small molecule. 204 genes were significantly down expressed and 55 genes were significantly up expressed in biofilms grown with 7µm/ml pyrimidine-diones small molecule. Among the 204 down expressed genes, 45 were hypothetical protein encoding gene and 159 were functional protein encoding genes. Of 55 up-regulated genes 21 were hypothetical genes and 34 were functional protein encoding genes. The functional annotation showed that gene involve in fatty acid metabolism, cell division, cell cycles, DNA metabolism, cell assembly were significantly down regulated and galactose metabolism related gene were up-expressed in biofilm grown with pyrimidine-diones small molecule.

Dataset Information

Structure of MurF from Streptococcus pneumoniae co-crystallized with a small molecule inhibitor exhibits interdomain closure.

Publications

Structure of MurF from Streptococcus pneumoniae co-crystallized with a small molecule inhibitor exhibits interdomain closure.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure