Evaluation of Two Models for Human Topoisomerase I Interaction with dsDNA and Camptothecin Derivatives

Human topoisomerase I (Top1) relaxes supercoiled DNA during cell division. Camptothecin stabilizes Top1/dsDNA covalent complexes which ultimately results in cell death, and this makes Top1 an anti-cancer target. There are two current models for how camptothecin and derivatives bind to Top1/dsDNA covalent complexes (Staker, et al., 2002, Proc Natl Acad Sci USA 99: 15387–15392; and Laco, et al., 2004, Bioorg Med Chem 12: 5225–5235). The interaction energies between bound camptothecin, and derivatives, and Top1/dsDNA in the two models were calculated. The published structure-activity-relationships for camptothecin and derivatives correlated with the interaction energies for camptothecin and derivatives in the Laco et al. model, however, this was not the case for several camptothecin derivatives in the Stacker et al. model. By defining the binding orientation of camptothecin and derivatives in the Top1/dsDNA active-site these results allow for the rational design of potentially more efficacious camptothecin derivatives

Development of Derivatives of 3, 3′-Diindolylmethane as Potent Leishmania donovani Bi-Subunit Topoisomerase IB Poisons

Background: The development of 3, 39-diindolyl methane (DIM) resistant parasite Leishmania donovani (LdDR50) by adaptation with increasing concentrations of the drug generates random mutations in the large and small subunits of heterodimeric DNA topoisomerase I of Leishmania (LdTOP1LS). Mutation of large subunit of LdTOP1LS at F270L is responsible for resistance to DIM up to 50 mM concentration. Methodology/Principal Findings: In search of compounds that inhibit the growth of the DIM resistant parasite and inhibit the catalytic activity of mutated topoisomerase I (F270L), we have prepared three derivatives of DIM namely DPDIM (2,29diphenyl 3,39-diindolyl methane), DMDIM (2,29-dimethyl 3,39-diindolyl methane) and DMODIM (5,59-dimethoxy 3,39diindolyl methane) from parent compound DIM. All the compounds inhibit the growth of DIM resistant parasites, induce DNA fragmentation and stabilize topo1-DNA cleavable complex with the wild type and mutant enzyme. Conclusion: The results suggest that the three derivatives of DIM can act as promising lead molecules for the generation of new anti-leishmanial agents

Structure of the cystathionine γ-synthase MetB from Mycobacterium ulcerans

Cystathionine γ-synthase (CGS) is a transferase that catalyzes the reaction between O 4-succinyl-l-homoserine and l-cysteine to produce l-­cystathionine and succinate. The crystal structure of CGS from M. ulcerans is presented covalently linked to the cofactor pyridoxal phosphate (PLP). A second structure contains PLP as well as a highly ordered HEPES molecule in the active site acting as a pseudo-ligand. This is the first structure ever reported from the pathogen M. ulcerans

Probing conformational states of glutaryl-CoA dehydrogenase by fragment screening

The first crystal structure is reported of a glutaryl-CoA dehydrogenase in the apo state without flavin adenine dinucleotide cofactor bound. Additional structures with small molecules complexed in the catalytic active site were obtained by fragment-based screening

Structure of 3-ketoacyl-(acyl-carrier-protein) reductase from Rickettsia prowazekii at 2.25 Å resolution

The R. prowazekii 3-ketoacyl-(acyl-carrier-protein) reductase is similar to those from other prokaryotic pathogens but differs significantly from the mammalian orthologue, strengthening its case as a potential drug target

BrabA.11339.a: anomalous diffraction and ligand binding guide towards the elucidation of the function of a ‘putative β-lactamase-like protein’ from Brucella melitensis

The structure of a β-lactamase-like protein from B. melitensis was solved independently using two data sets with anomalous signal. Anomalous Fourier maps could confirm the identity of two metal ions in the active site. AMP-bound and GMP-bound structures provide hints to the possible function of the protein

Inhibitor-bound complexes of dihydrofolate reductase-thymidylate synthase from Babesia bovis

Structural characterization of the bifunctional enzyme dihydrofolate reductase-thymidylate synthase from B. bovis in the apo state and complexed with antifolate inhibitors in both enzymatic active sites is reported

Structure of a Nudix hydrolase (MutT) in the Mg2+-­bound state from Bartonella henselae, the bacterium responsible for cat scratch fever

B. henselae is the etiological agent responsible for cat scratch fever (bartonellosis). The crystal structure of the smaller of the two Nudix hydrolases encoded in the genome of B. henselae, Bh-MutT, was determined to 2.1 Å resolution

The Protein Maker: an automated system for high-throughput parallel purification

The Protein Maker instrument addresses a critical bottleneck in structural genomics by allowing automated purification and buffer testing of multiple protein targets in parallel with a single instrument. Here, the use of this instrument to (i) purify multiple influenza-virus proteins in parallel for crystallization trials and (ii) identify optimal lysis-buffer conditions prior to large-scale protein purification is described