Crystal structure of ferrioxamine B: a comparative analysis and implications for molecular recognition

Ferrioxamine B was successfully co-crystallized with ethanolpentaaquomagnesium(II) and perchlorate ions as counter ions, C27H62Cl3FeMgN6O26, and the crystal structure has been determined by single-crystal X-ray diffraction. The crystals are monoclinic, space group P2(1)/n, four molecules per unit cell with dimensions a=21.1945(7) A, b=10.0034(3) A, c=106.560(1) A, and beta=106.560(1) degrees. The crystal structure contains a racemic mixture of Lambda-N-cis,cis and Delta-N-cis,cis coordination isomers. The structural parameters and the conformational features of ferrioxamine B compare very well with those of ferrioxamines D1 and E, with an exception of the orientation of the pendant protonated amine, which is pointing away from the connecting amide chains and towards the carbonyl face of the inner coordination shell distorted octahedron. This pendant protonated amine, in conjunction with the carbonyl face of the Fe(III) coordination shell, is proposed to play an important role in the recognition and membrane transport processes.     

Structure-based optimization of 1H-imidazole-2-carboxamides as Axl kinase inhibitors utilizing a Mer mutant surrogate

Axl has been a target of interest in the oncology field for several years based on its role in various oncogenic processes. To date, no wild-type Axl crystal structure has been reported. Herein, we describe the structure-based optimization of a novel chemotype of Axl inhibitors, 1H-imidazole-2-carboxamide, using a mutated kinase homolog, Mer(I650M), as a crystallographic surrogate. Iterative optimization of the initial lead compound (1) led to compound (21), a selective and potent inhibitor of wild-type Axl. Compound (21) will serve as a useful compound for further in vivo studies.     

Ameloblastin peptide encoded by exon 5 interacts with amelogenin N-terminus

Interactions between enamel matrix proteins are important for enamel biomineralization. In recent in situ studies, we showed that the N-terminal proteolytic product of ameloblastin co-localized with amelogenin around the prism boundaries. However, the molecular mechanisms of such interactions are still unclear. Here, in order to determine the interacting domains between amelogenin and ameloblastin, we designed four ameloblastin peptides derived from different regions of the full-length protein (AB1, AB2 and AB3 at N-terminus, and AB6 at C-terminus) and studied their interactions with recombinant amelogenin (rP172), and the tyrosine-rich amelogenin polypeptide (TRAP). A series of amelogenin Trp variants (rP172(W25), rP172(W45) and rP172(W161)) were also used for intrinsic fluorescence spectroscopy. Fluorescence spectra of rP172 titrated with AB3, a peptide encoded by exon 5 of ameloblastin, showed a shift in λ_max in a dose-dependent manner, indicating molecular interactions in the region encoded by exon 5 of ameloblastin. Circular dichroism (CD) spectra of amelogenin titrated with AB3 showed that amelogenin was responsible for forming α-helix in the presence of ameloblastin. Fluorescence spectra of amelogenin Trp variants as well as the spectra of TRAP titrated with AB3 showed that the N-terminus of amelogenin is involved in the interaction between ameloblastin and amelogenin. We suggest that macromolecular co-assembly between amelogenin and ameloblastin may play important roles in enamel biomineralization.     

Protective effect of Premna tomentosa extract (L. verbanacae) on acetaminophen-induced mitochondrial dysfunction in rats

Allurement of herbs as health beneficial foods (physiologically functional foods) and as a source material for the development of new drugs, has led to greater furtherance in the study of herbal medicines during recent years. Plant extracts are being utilized to treat a wide variety of diseases like hepatotoxicity. Premna tomentosa is one such medicinal plant used widely in Indian ayurvedic medicine for the treatment of liver disorders. This study appraised the effectiveness of P. tomentosa leaf extract in protecting the liver against mitochondrial damage induced by acetaminophen, since mitochondrial injury has been investigated as a potential initiator of hepatotoxicity. Normal Wistar strain rats were pre-treated with P. tomentosa extract (750 mg/kg, orally) for 15 days and then intoxicated with acetaminophen (640 mg/kg, orally). Mitochondria were isolated from liver of experimental animals and assessed for the levels of lipid peroxide products, GSH and mitochondrial enzymes (isocitrate dehydrogenase, -keto glutarate dehydrogenase, succinate dehydrogenase, malate dehydrogenase, NADH dehydrogenase and cytochrome-C-oxidase). The levels of Lipid peroxidation products were increased and the levels of the other assessed parameters were significantly decreased in hepatotoxicity induced animals. Whereas, the levels were brought back to normal in P. tomentosa pre-treated rats, which shows the protective effect of the extract against mitochondrial damage. Presence of anti-oxidant compound d-limonene (58%) in P. tomentosa leaves, which is known to enhance conjugation of toxic metabolites by maintaining liver GSH concentrations may explain the hepatoprotective property of the extract.     

Double-spanning plant viral movement protein integration into the endoplasmic reticulum membrane is signal recognition particle-dependent, translocon-mediated, and concerted

The current model for cell-to-cell movement of plant viruses holds that transport requires virus-encoded movement proteins that intimately associate with endoplasmic reticulum membranes. We have examined the early stages of the integration into endoplasmic reticulum membranes of a double-spanning viral movement protein using photocross-linking. We have discovered that this process is cotranslational and proceeds in a signal recognition particle-dependent manner. In addition, nascent chain photocross-linking to Sec61alpha and translocating chain-associated membrane protein reveal that viral membrane protein insertion takes place via the translocon, as with most eukaryotic membrane proteins, but that the two transmembrane segments of the viral protein leave the translocon and enter the lipid bilayer together.     

Kinetics of iron release from ferric binding protein (FbpA): mechanistic implications in bacterial periplasm-to-cytosol Fe3+ transport

The ferric binding protein (FbpA) transports iron across the periplasmic space of certain Gram-negative bacteria and is an important component involved in iron acquisition by pathogenic Neisseria spp. (Neisseria gonorrheae and Neisseria meningitidis). Previous work has demonstrated that the synergistic anion, required for tight Fe(3+) sequestration by FbpA, also plays a key role in inserting Fe(3+) into the FbpA binding site. Here, we investigate the iron release process from various forms of holo-FbpA, Fe(3+)FbpA-X, during the course of a chelator competition reaction using EDTA and Tiron. Fe(3+)FbpA-X represents the protein assembly complex with different synergistic anions, X = PO(4)(3)(-) and NTA. Stepwise mechanisms of Fe(3+) release are proposed on the basis of kinetic profiles of these chelator competition reactions. Fe(3+)FbpA-PO(4) and Fe(3+)FbpA-NTA react differently with EDTA and Tiron during the Fe(3+)-exchange process. EDTA replaces PO(4)(3)(-) and NTA from the first coordination shell of Fe(3+) and acts as a synergistic anion to give a spectroscopically distinguishable intermediate, Fe(3+)FbpA-EDTA, prior to pulling Fe(3+) out of the protein. Tiron, on the other hand, does not act as a synergistic anion but is a more efficient competing chelator as it removes Fe(3+) from FbpA at rate much faster than EDTA. These results reaffirm the contribution of the synergistic anion to the FbpA iron transport process as the anion, in addition to playing a facilitative role in iron binding, appears to have a "gatekeeper" role, thereby modulating the Fe(3+) release process.     

Using workflows to explore and optimise named entity recognition for chemistry

Chemistry text mining tools should be interoperable and adaptable regardless of system-level implementation, installation or even programming issues. We aim to abstract the functionality of these tools from the underlying implementation via reconfigurable workflows for automatically identifying chemical names. To achieve this, we refactored an established named entity recogniser (in the chemistry domain), OSCAR and studied the impact of each component on the net performance. We developed two reconfigurable workflows from OSCAR using an interoperable text mining framework, U-Compare. These workflows can be altered using the drag-&-drop mechanism of the graphical user interface of U-Compare. These workflows also provide a platform to study the relationship between text mining components such as tokenisation and named entity recognition (using maximum entropy Markov model (MEMM) and pattern recognition based classifiers). Results indicate that, for chemistry in particular, eliminating noise generated by tokenisation techniques lead to a slightly better performance than others, in terms of named entity recognition (NER) accuracy. Poor tokenisation translates into poorer input to the classifier components which in turn leads to an increase in Type I or Type II errors, thus, lowering the overall performance. On the Sciborg corpus, the workflow based system, which uses a new tokeniser whilst retaining the same MEMM component, increases the F-score from 82.35% to 84.44%. On the PubMed corpus, it recorded an F-score of 84.84% as against 84.23% by OSCAR.     

Microbial transformation of a fungicide-derived amine, dimethyl-p-phenylene diamine by Alcaligenes DM4

Abstract Microbial transformation of N , N -dimethyl- p -phenylene diamine (DMPDA), a microbial product formed from the fungicide fenaminosulf ( p -dimethylaminobenzenediazo sodium sulfonate) was studied by enriching microbes in soils treated with the amine. Microorganisms isolated from DMPDA-treated soil belonged to the genera of Micrococcus, Alcaligenes , and Corynebacterium . Of the various isolates, Alcaligenes DM₄ showed maximal growth on DMPDA utilizing it as sources of carbon and nitrogen. When grown in mineral salts basal medium containing 0.05% DMPDA to serve as carbon and nitrogen sources, Alcaligenes DM₄ grew exponentially up to 18 h. Even though the characterization of the complete pathway of microbial degradation of DMPDA could not be carried out due to the auto-oxidation of the compound, the initial transformation product of DMPDA by Alcaligenes DM₄ has been identified as a dimer. The dimer is generated into the culture medium presumably by the extra-cellular oxidase of Alcaligenes DM₄. It is suggested that the risk-benefit evaluation on the use of fenaminosulf is to be made taking into consideration the microbial transformations of the fungicide.     

Structural adaptation of tooth enamel protein amelogenin in the presence of SDS micelles

Amelogenin, the major extracellular matrix protein of developing tooth enamel is intrinsically disordered. Through its interaction with other proteins and mineral, amelogenin assists enamel biomineralization by controlling the formation of highly organized enamel crystal arrays. We used circular dichroism (CD), dynamic light scattering (DLS), fluorescence, and NMR spectroscopy to investigate the folding propensity of recombinant porcine amelogenin rP172 following its interaction with SDS, at levels above critical micelle concentration. The rP172‐SDS complex formation was confirmed by DLS, while an increase in the structure moiety of rP172 was noted through CD and fluorescence experiments. Fluorescence quenching analyses performed on several rP172 mutants where all but one Trp was replaced by Tyr at different sequence regions confirmed that the interaction of amelogenin with SDS micelles occurs via the N‐terminal region close to Trp25 where helical segments can be detected by NMR. NMR spectroscopy and structural refinement calculations using CS‐Rosetta modeling confirm that the highly conserved N‐terminal domain is prone to form helical structure when bound to SDS micelles. Our findings reported here reveal interactions leading to significant changes in the secondary structure of rP172 upon treatment with SDS. These interactions may reflect the physiological relevance of the flexible nature of amelogenin and its sequence specific helical propensity that might enable it to structurally adapt with charged and potential targets such as cell surface, mineral, and other proteins during enamel biomineralization. © 2013 Wiley Periodicals, Inc. Biopolymers 101: 525–535, 2014.