Flexibility and inhibitor binding in Cdc25 phosphatases

Cdc25 phosphatases involved in cell cycle checkpoints are now active targets for the development of anti-cancer therapies. Rational drug design would certainly benefit from detailed structural information for Cdc25s. However, only apo- or sulfate-bound crystal structures of the Cdc25 catalytic domain have been described so far. Together with previously available crystalographic data, results from molecular dynamics simulations, bioinformatic analysis, and computer-generated conformational ensembles shown here indicate that the last 30-40 residues in the C-terminus of Cdc25B are partially unfolded or disordered in solution. The effect of C-terminal flexibility upon binding of two potent small molecule inhibitors to Cdc25B is then analyzed by using three structural models with variable levels of flexibility, including an equilibrium distributed ensemble of Cdc25B backbone conformations. The three Cdc25B structural models are used in combination with flexible docking, clustering, and calculation of binding free energies by the linear interaction energy approximation to construct and validate Cdc25B-inhibitor complexes. Two binding sites are identified on top and beside the Cdc25B active site. The diversity of interaction modes found increases with receptor flexibility. Backbone flexibility allows the formation of transient cavities or compact hydrophobic units on the surface of the stable, folded protein core that are unexposed or unavailable for ligand binding in rigid and densely packed crystal structures. The present results may help to speculate on the mechanisms of small molecule complexation to partially unfolded or locally disordered proteins.

Ixolaris, a tissue factor inhibitor, blocks primary tumor growth and angiogenesis in a glioblastoma model

Background: The expression levels of the clotting initiator protein Tissue Factor (TF) correlate with vessel density and the histological malignancy grade of glioma patients. Increased procoagulant tonus in high grade tumors (glioblastomas) also indicates a potential role for TF in progression of this disease, and suggests that anticoagulants could be used as adjuvants for its treatment. Objectives: We hypothesized that blocking of TF activity with the tick anticoagulant Ixolaris might interfere with glioblastoma progression. Methods and results: TF was identified in U87-MG cells by flow-cytometric and functional assays (extrinsic tenase). In addition, flow-cytometric analysis demonstrated the exposure of phosphatidylserine in the surface of U87-MG cells, which supported the assembly of intrinsic tenase (FIXa/FVIIIa/FX) and prothrombinase (FVa/FXa/prothrombin) complexes, accounting for the production of FXa and thrombin, respectively. Ixolaris effectively blocked the in vitro TF-dependent procoagulant activity of the U87-MG human glioblastoma cell line and attenuated multimolecular coagulation complexes assembly. Notably, Ixolaris inhibited the in vivo tumorigenic potential of U87-MG cells in nude mice, without observable bleeding. This inhibitory effect of Ixolaris on tumor growth was associated with downregulation of VEGF and reduced tumor vascularization. Conclusion: Our results suggest that Ixolaris might be a promising agent for anti-tumor therapy in humans.

Immobilized analogues of sunflower trypsin inhibitor-1 constitute a versatile group of affinity sorbents for selective isolation of serine proteases

Sunflower trypsin inhibitor-1 (SFI-1), a natural 14-residue cyclic peptide, and some of its synthetic acyclic variants are potent protease inhibitors displaying peculiar inhibitory profiles. Here we describe the synthesis and use of affinity sorbents prepared by coupling SFTI-1 analogues to agarose resin. Chymotrypsinand trypsin-like proteases could then be selectively isolated from pancreatin; similarly, other proteases were obtained from distinct biological sources. The binding capacity of [Lys5]-SFTI-1-agarose for trypsin was estimated at over 10 mg/mL of packed gel. SFTI-1-based resins could find application either to improve the performance of current purification protocols or as novel protease-discovery tools in different areas of biological investigation. (C) 2009 Elsevier B.V. All rights reserved.

Effect of trypsin inhibitor from Crotalaria pallida seeds on Callosobruchus maculatus (cowpea weevil) and Ceratitis capitata (fruit fly)

GOMES, Carlos E. M. et al. Effect of trypsin inhibitor from Crotalaria pallida seeds on Callosobruchus maculatus (cowpea weevil) and Ceratitis capitata (fruit fly). Plant Physiology and Biochemistry (Paris), v. 43, n. 12, p. 1095-1102, 2005.ISSN 0981-9428. DOI:10.1016/j.plaphy.2005.11.004.

Quercetin as an inhibitor of snake venom secretory phospholipase A2

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Second generation European corn borer injury and Irish potato physiology, yield, and quality

European corn borer (ECB) [Ostrinia nubilalis (Hubner)] (Lepidoptera: Crambidae) is known to infest Irish potato (Solanum tuberosum L.) but only causes economic damage during the first generation in East Coast potato producing areas. However, in Nebraska, second generation ECB infest potato plants during the bulking period and may reduce yield and/or potato quality. Experiments were conducted in 2001, 2002, and 2003 to examine physiological and yield effects of second generation ECB injury to potato in Nebraska. Pike, Atlantic, and three Frito Lay proprietary varieties (FL1867, FL1879, and FL1833) were used. Experimental plots were infested with four ECB egg masses per plant to simulate ECB infestation by second-generation larvae; controls received no egg masses. Photosynthetic rates, tuber weights, tuber size grades, solids, and fry quality were measured. Potato plants with ECB infestation had significantly reduced photosynthetic rates on ECB-infested stems and on uninfested stems on the same plant when larvae were in the fifth instar. When insects were in the fourth instar, photosynthetic rates were reduced only on ECB-infested stems. In 2001, ECB infestation reduced the average mass of large tubers and increased the amount of small tubers in FL1867 and FL1879. In 2002, significant yield reductions were not observed. Across both years, ECB-infested plots produced fewer large (65- to 100-mm diam.) tubers than control plots. Other tuber properties and chip qualities were unaffected. This study indicates that second generation ECB infestation of approximately 30% infested plants results in economic loss for some chipping varieties and affects tuber bulking. In contrast to east coast growers, Midwest potato farmers must be concerned with second generation ECB.

Effects of Silicon and Drought Stress on Tuber Yield and Leaf Biochemical Characteristics in Potato

Silicon has beneficial effects on many crops, mainly under biotic and abiotic stresses. Silicon can affect biochemical, physiological, and photosynthetic processes and, consequently, alleviates drought stress. However, the effects of Si on potato (Solanum tuberosum L.) plants under drought stress are still unknown. The objective of this study was to evaluate the effect of Si supply on some biochemical characteristics and yield of potato tubers, either exposed or not exposed to drought stress. The experiment was conducted in pots containing 50 dm(3) of a Typic Acrortox soil (33% clay, 4% silt, and 63% sand). The treatments consisted of the absence or presence of Si application (0 and 284.4 mg dm(-3)), through soil amelioration with dolomitic lime and Ca and Mg silicate, and in the absence or presence of water deficit (-0.020 MPa and -0.050 MPa soil water potential, respectively), with eight replications. Silicon application and water deficit resulted in the greatest Si concentration in potato leaves. Proline concentrations increased under lower water availability and higher Si availability in the soil, which indicates that Si may be associated with plant osmotic adjustment. Water deficit and Si application decreased total sugars and soluble proteins concentrations in the leaves. Silicon application reduced stalk lodging and increased mean tuber weight and, consequently, tuber yield, especially in the absence of water stress.

Agronomic characteristics, isoflavone content and Kunitz trypsin inhibitor of vegetable soybean genotypes

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Nutrition, dry matter accumulation and partitioning and phosphorus use efficiency of potato grown at different phosphorus levels in nutrient solution

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

The field resistance of potato cultivars to foliar late blight and its relationship with foliage maturity type and skin type in Brazil

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

A tellurium-based cathepsin B inhibitor: Molecular structure, modelling, molecular docking and biological evaluation

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Highly polymorphic in silico-derived microsatellite loci in the potato-infecting fungal pathogen Rhizoctonia solani anastomosis group 3 from the Colombian Andes

Fourteen polymorphic microsatellite DNA markers derived from the draft genome sequence of Rhizoctonia solani anastomosis group 3 (AG-3), strain Rhs 1AP, were designed and characterized from the potato-infecting soil fungus R. solani AG-3. All loci were polymorphic in two field populations collected from Solanum tuberosum and S. phureja in the Colombian Andes. The total number of alleles per locus ranged from two to seven, while gene diversity (expected heterozygosity) varied from 0.11 to 0.81. Considering the variable levels of genetic diversity observed, these markers should be useful for population genetic analyses of this important dikaryotic fungal pathogen on a global scale.

Protective effects of Tacrolimus, a calcineurin inhibitor, in experimental periodontitis in rats

Objective: Periodontitis is a well-appreciated example of leukocyte-mediated bone loss and inflammation with pathogenic features similar to those observed in other inflammatory diseases, such as arthritis. Since Tacrolimus, is an immunomodulatory drug used for the treatment of some cases of arthritis, we hypothesized that it may modulate periodontal disease.Design: Using a murine model of ligature-induced periodontal disease, we assessed the effects of daily administrations of Tacrolimus (1 mg/kg body weight) on bone loss, enzymatic (myeloperoxidase) analysis, differential white blood cells counts, airpouch exudate and cytokine expression for 5-30 days.Results: Radiographic, enzymatic (myeloperoxidase) and histological analysis revealed that Tacrolimus reduced the severity of periodontitis. More specifically, Tacrolimus suppressed the expression of serum interleukin (IL-1 beta), tumour necrosis factor (TNF-alpha), IL-6, airpouch exudate PGE(2) and leukocytosis usually observed after the induction of periodontitis. Tacrolimus treatment in periodontitis-induced rats conferred protection against the inflammation-induced tissue and bone loss associated with periodontitis, through a mechanism involving IL-1 beta, TNF-alpha and IL-6.Conclusions: the effects of Tacrolimus on periodontal disease pathogenesis may provide clues to a novel approach to host modulation therapy in destructive periodontal disease. (C) 2007 Elsevier Ltd. All rights reserved.