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221.
Trosset JY Dalvit C Knapp S Fasolini M Veronesi M Mantegani S Gianellini LM Catana C Sundström M Stouten PF Moll JK 《Proteins》2006,64(1):60-67
The interaction between beta-catenin and Tcf family members is crucial for the Wnt signal transduction pathway, which is commonly mutated in cancer. This interaction extends over a very large surface area (4800 A(2)), and inhibiting such interactions using low molecular weight inhibitors is a challenge. However, protein surfaces frequently contain "hot spots," small patches that are the main mediators of binding affinity. By making tight interactions with a hot spot, a small molecule can compete with a protein. The Tcf3/Tcf4-binding surface on beta-catenin contains a well-defined hot spot around residues K435 and R469. A 17,700 compounds subset of the Pharmacia corporate collection was docked to this hot spot with the QXP program; 22 of the best scoring compounds were put into a biophysical (NMR and ITC) screening funnel, where specific binding to beta-catenin, competition with Tcf4 and finally binding constants were determined. This process led to the discovery of three druglike, low molecular weight Tcf4-competitive compounds with the tightest binder having a K(D) of 450 nM. Our approach can be used in several situations (e.g., when selecting compounds from external collections, when no biochemical functional assay is available, or when no HTS is envisioned), and it may be generally applicable to the identification of inhibitors of protein-protein interactions. 相似文献
222.
Carbone L Nergadze SG Magnani E Misceo D Francesca Cardone M Roberto R Bertoni L Attolini C Francesca Piras M de Jong P Raudsepp T Chowdhary BP Guérin G Archidiacono N Rocchi M Giulotto E 《Genomics》2006,87(6):777-782
Centromere repositioning (CR) is a recently discovered biological phenomenon consisting of the emergence of a new centromere along a chromosome and the inactivation of the old one. After a CR, the primary constriction and the centromeric function are localized in a new position while the order of physical markers on the chromosome remains unchanged. These events profoundly affect chromosomal architecture. Since horses, asses, and zebras, whose evolutionary divergence is relatively recent, show remarkable morphological similarity and capacity to interbreed despite their chromosomes differing considerably, we investigated the role of CR in the karyotype evolution of the genus Equus. Using appropriate panels of BAC clones in FISH experiments, we compared the centromere position and marker order arrangement among orthologous chromosomes of Burchelli's zebra (Equus burchelli), donkey (Equus asinus), and horse (Equus caballus). Surprisingly, at least eight CRs took place during the evolution of this genus. Even more surprisingly, five cases of CR have occurred in the donkey after its divergence from zebra, that is, in a very short evolutionary time (approximately 1 million years).These findings suggest that in some species the CR phenomenon could have played an important role in karyotype shaping, with potential consequences on population dynamics and speciation. 相似文献
223.
Heirman I Ginneberge D Brigelius-Flohé R Hendrickx N Agostinis P Brouckaert P Rottiers P Grooten J 《Free radical biology & medicine》2006,40(2):285-294
Using tumor cell-restricted overexpression of glutathione peroxidase 4 (GP x 4), we investigated the contribution of tumor cell eicosanoids to solid tumor growth and malignant progression in two tumor models differing in tumorigenic potential. By lowering cellular lipid hydroperoxide levels, GP x 4 inhibits cyclooxygenase (COX) and lipoxygenase (LOX) activities. GP x 4 overexpression drastically impeded solid tumor growth of weakly tumorigenic L929 fibrosarcoma cells, whereas B16BL6 melanoma solid tumor growth was unaffected. Yet, GP x 4 overexpression did markedly increase the sensitivity of B16BL6 tumors to angio-destructive TNF-alpha therapy and abolished the metastatic lung colonizing capacity of B16BL6 cells. Furthermore, the GP x 4-mediated suppression of tumor cell prostaglandin E(2) (PGE(2)) production impeded the induction of COX-2 expression by the tumor stress conditions hypoxia and inflammation. Thus, our results reflect a PGE(2)-driven positive feedback loop for COX-2 expression in tumor cells. This was further supported by the restoration of COX-2 induction capacity of GP x 4-overexpressing L929 tumor cells when cultured in the presence of exogenous PGE(2). Thus, although COX-2 expression and eicosanoid production may be enabled by PGE(2) from the tumor microenvironment, our results demonstrate the predominant tumor cell origin of protumoral eicosanoids, promoting solid tumor growth of weakly tumorigenic tumors and malignant progression of strongly tumorigenic tumors. 相似文献
224.
225.
Pieter?PypersEmail author Josefien?Delrue Jan?Diels Erik?Smolders Roel?Merckx 《Plant and Soil》2006,284(1-2):217-227
Phosphorus (P) uptake by plant roots depends on P intensity (I) and P quantity (Q) in the soil. The relative importance of Q and I on P uptake is unknown for soils with large P sorption capacities because of difficulties in determining trace levels of
P in the soil solution. We applied a new isotope based method to detect low P concentrations (<20 μg P l−1). The Q factor was determined by assessment of the isotopically exchangeable P in the soil (E-value) and the I factor was determined by measurement of the P concentration in the pore water. A pot trial was set up using four soils with
similar labile P quantities but contrasting P buffering capacities. Soils were amended with KH2PO4 at various rates and pigeon pea (Cajanus cajan L.) was grown for 25 days. The P intensity ranged between 0.0008 and 50 mg P l−1 and the P quantity ranged between 10 and 500 mg P kg−1. Shoot dry matter (DM) yield and P uptake significantly increased with increasing P application rates in all soils. Shoot
DM yield and P uptake, relative to the maximal yield or P uptake, were better correlated with the P concentration in the pore
water (R
2 = 0.83–0.90) than with the E-value (R
2=0.40–0.53). The observed P uptakes were strongly correlated to values simulated using a mechanistic rhizosphere model (NST
3.0). A sensitivity analysis reveals that the effect of P intensity on the short-term P uptake by pigeon pea exceeded the
effect of P quantity both at low and high P levels. However, DM yield and P uptake at a given P intensity consistently increased
with increasing P buffering capacity (PBC). The experimental data showed that the intensity yielding 80% of the maximal P
uptake was 4 times larger in the soil with the smallest PBC compared to the soil with the largest PBC. This study confirms
that short-term P uptake by legumes is principally controlled by the P intensity in the soil, but is to a large extent also
affected by the PBC of the soil.
Section Editor: N. J. Barrow 相似文献
226.
Moons P Van Houdt R Aertsen A Vanoirbeek K Engelborghs Y Michiels CW 《Applied and environmental microbiology》2006,72(11):7294-7300
We have previously characterized the N-acyl-l-homoserine lactone-based quorum-sensing system of the biofilm isolate Serratia plymuthica RVH1. Here we investigated the role of quorum sensing and of quorum-sensing-dependent production of an antimicrobial compound (AC) on biofilm formation by RVH1 and on the cocultivation of RVH1 and Escherichia coli in planktonic cultures or in biofilms. Biofilm formation of S. plymuthica was not affected by the knockout of splI or splR, the S. plymuthica homologs of the luxI or luxR quorum-sensing gene, respectively, or by the knockout of AC production. E. coli grew well in mixed broth culture with RVH1 until the latter reached 8.5 to 9.5 log CFU/ml, after which the E. coli colony counts steeply declined. In comparison, only a very small decline occurred in cocultures with the S. plymuthica AC-deficient and splI mutants. Complementation with exogenous N-hexanoyl-l-homoserine lactone rescued the wild-type phenotype of the splI mutant. The splR knockout mutant also induced a steep decline of E. coli, consistent with its proposed function as a repressor of quorum-sensing-regulated genes. The numbers of E. coli in 3-day-old mixed biofilms followed a similar pattern, being higher with S. plymuthica deficient in SplI or AC production than with wild-type S. plymuthica, the splR mutant, or the splI mutant in the presence of N-hexanoyl-l-homoserine lactone. Confocal laser scanning microscopic analysis of mixed biofilms established with strains producing different fluorescent proteins showed that E. coli microcolonies were less developed in the presence of RVH1 than in the presence of the AC-deficient mutant. 相似文献
227.
Termont S Vandenbroucke K Iserentant D Neirynck S Steidler L Remaut E Rottiers P 《Applied and environmental microbiology》2006,72(12):7694-7700
Interleukin-10 (IL-10) is a promising candidate for the treatment of inflammatory bowel disease. Intragastric administration of Lactococcus lactis genetically modified to secrete IL-10 in situ in the intestine was shown to be effective in healing and preventing chronic colitis in mice. However, its use in humans is hindered by the sensitivity of L. lactis to freeze-drying and its poor survival in the gastrointestinal tract. We expressed the trehalose synthesizing genes from Escherichia coli under control of the nisin-inducible promoter in L. lactis. Induced cells accumulated intracellular trehalose and retained nearly 100% viability after freeze-drying, together with a markedly prolonged shelf life. Remarkably, cells producing trehalose were resistant to bile, and their viability in human gastric juice was enhanced. None of these effects were seen with exogenously added trehalose. Trehalose accumulation did not interfere with IL-10 secretion or with therapeutic efficacy in murine colitis. The newly acquired properties should enable a larger proportion of the administered bacteria to reach the gastrointestinal tract in a bioactive form, providing a means for more effective mucosal delivery of therapeutics. 相似文献
228.
Dorrestein PC Blackhall J Straight PD Fischbach MA Garneau-Tsodikova S Edwards DJ McLaughlin S Lin M Gerwick WH Kolter R Walsh CT Kelleher NL 《Biochemistry》2006,45(6):1537-1546
For screening a pool of potential substrates that load carrier domains found in nonribosomal peptide synthetases, large molecule mass spectrometry is shown to be a new, unbiased assay. Combining the high resolving power of Fourier transform mass spectrometry with the ability of adenylation domains to select their own substrates, the mass change that takes place upon formation of a covalent intermediate thus identifies the substrate. This assay has an advantage over traditional radiochemical assays in that many substrates, the substrate pool, can be screened simultaneously. Using proteins on the nikkomycin, clorobiocin, coumermycin A1, yersiniabactin, pyochelin, and enterobactin biosynthetic pathways as proof of principle, preferred substrates are readily identified from substrate pools. Furthermore, this assay can be used to provide insight into the timing of tailoring events of biosynthetic pathways as demonstrated using the bromination reaction found on the jamaicamide biosynthetic pathway. Finally, this assay can provide insight into the role and function of orphan gene clusters for which the encoded natural product is unknown. This is demonstrated by identifying the substrates for two NRPS modules from the pksN and pksJ genes that are found on an orphan NRPS/PKS hybrid cluster from Bacillus subtilis. This new assay format is especially timely for activity screening in an era when new types of thiotemplate assembly lines that defy classification are being discovered at an accelerating rate. 相似文献
229.
Angelika Lehner Sabine Nitzsche Pieter Breeuwer Benjamin Diep Karin Thelen Roger Stephan 《BMC microbiology》2006,6(1):15-8
Background
Enterobacter sakazakii is a foodborne pathogen that has been associated with sporadic cases and outbreaks causing meningitis, necrotizing enterocolitis and sepsis especially in neonates. The current FDA detection method includes two enrichment steps, the subculturing of the second enrichment broth on a selective agar (VRBG), a further subculturing of selected grown colonies on TSA and the subsequent biochemical identification of yellow-pigmented colonies by API20E. However, there is a strong need for simplified methods for isolation and identification of E. sakazakii. In this study, two chromogenic media, which allow to indicate presumptive E. sakazakii colonies by the alpha glucosidase activity, as well as a newly developed 1,6-alpha-glucosidase based conventional PCR assay and a rRNA oligonucleotide probe based commercial test system for identification of presumptive E. sakazakii were evaluated on 98 target and non-target strains. The methods were compared with respect to specifiCity aspects. 相似文献230.
Bruggeman V Van As P Decuypere E 《Comparative biochemistry and physiology. Part A, Molecular & integrative physiology》2002,131(4):839-846
In mammals, the phenotype of the homogametic sex develops in the (relative) absence of steroids and the phenotype of the heterogametic sex is imposed by the early action of steroids. In contrast, the heterogametic sex in avian species is the female and the presence of estrogens and their receptors plays a crucial role in female sexual differentiation. The time- and sex-dependent expression of enzymes involved in steroidogenesis which determine the ratio of androgens/estrogens produced by the gonads has been extensively investigated during the last 5-6 years. These results all show that the lack of estrogen synthesis in the male appears to be due to the extremely low levels of 17beta-hydroxysteroid dehydrogenase and P450aromatase expression. In females, extensive expression of the aromatase gene (around day 5-6 of incubation), leading to estrogen synthesis, and specific expression of the estrogen receptor-mRNA in the left gonad results in the development of a functional left ovary. Other sex differences can be found in the expression of the inhibin subunit genes in gonads of chicken embryos and in circulating concentrations of inhibin, follicle stimulating hormone (FSH) and steroids. Sex reversal attempts have been made by varying incubation temperatures, by using anti-estrogens, androgens, aromatase inhibitors and synthetic steroids. In ovo administration of a sex steroid hormone or an inhibitor of endogenous sex steroid synthesis can cause phenotypical sex reversal. All these experiments show that the development of gonads in birds is very sensitive to changes in the embryonic hormonal environment, sometimes resulting in changes of postnatal reproduction and even growth. 相似文献