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排序方式: 共有539条查询结果,搜索用时 31 毫秒
121.
【目的】本研究以根癌农杆菌C58为材料,鉴定其甲基趋化受体蛋白(methyl-accepting chemotaxis protein,MCP) MCP1912能够识别的配体,并研究该蛋白在调控根癌农杆菌趋化响应中的具体功能。【方法】通过异源表达MCP1912的配体结合结构域(ligand binding domain,LBD),获得带有His标签的LBD蛋白(LBD1912)。利用基于荧光的热位移测定法(fluorescence-based thermal shift assay,TSA)筛选出LBD1912的潜在配体;通过等温滴定量热(isothermal titration calorimetry,ITC),进一步确定筛选出的潜在配体,并测定LBD1912与配体结合之后的解离平衡常数KD。利用基于同源重组的精准DNA片段删除方法,敲除根癌农杆菌C58中编码MCP1912的基因atu1912,获得MCP1 912 相似文献
122.
Sean R Collins Hee Won Yang Kimberly M Bonger Emmanuel G Guignet Thomas J Wandless Tobias Meyer 《Molecular systems biology》2015,11(4)
Numerous molecular components have been identified that regulate the directed migration of eukaryotic cells toward sources of chemoattractant. However, how the components of this system are wired together to coordinate multiple aspects of the response, such as directionality, speed, and sensitivity to stimulus, remains poorly understood. Here we developed a method to shape chemoattractant gradients optically and analyze cellular chemotaxis responses of hundreds of living cells per well in 96‐well format by measuring speed changes and directional accuracy. We then systematically characterized migration and chemotaxis phenotypes for 285 siRNA perturbations. A key finding was that the G‐protein Giα subunit selectively controls the direction of migration while the receptor and Gβ subunit proportionally control both speed and direction. Furthermore, we demonstrate that neutrophils chemotax persistently in response to gradients of fMLF but only transiently in response to gradients of ATP. The method we introduce is applicable for diverse chemical cues and systematic perturbations, can be used to measure multiple cell migration and signaling parameters, and is compatible with low‐ and high‐resolution fluorescence microscopy. 相似文献
123.
Sphingosine-1-phosphate signaling regulates lamellipodia localization of cortactin complexes in endothelial cells 总被引:2,自引:2,他引:0
Sphingosine-1-phosphate (S1P), a serum-borne lipid mediator, was demonstrated to be a potent chemoattractant of endothelial cells. It was recently shown that the colocalization of cortactin and actin related protein 2/3 (Arp2/3) in the lamellipodia is critical to S1P-induced endothelial chemotaxis. In this report, we describe that S1P-stimulated cortactin translocation to the cell periphery to form lamellipodia is specifically mediated by the endothelial S1P1 G-protein coupled receptor, and is regulated by Gi-mediated Akt-dependent S1P1 receptor phosphorylation and Cdc42/Rac activation pathways. In contrast to Src-dependent fibroblast growth factor-induced cortactin translocation, tyrosine phosphorylation cascades are not required for S1P-mediated lamellipodia formation and chemotaxis. Furthermore, we also demonstrate that S1P signaling, via the Gi/Akt/S1P1 phosphorylation/Rac pathway, regulates the cortactin–Arp2/3 complex formation, which ultimately results in membrane ruffling, formation of the lamellipodia and endothelial migration.J.F. Lee and H. Ozaki contributed equally to this work 相似文献
124.
Mizuno K Padma P Konno A Satouh Y Ogawa K Inaba K 《Biology of the cell / under the auspices of the European Cell Biology Organization》2009,101(2):91-103
Background information. Spermatozoa show several changes in flagellar waveform, such as upon fertilization. Ca2+ has been shown to play critical roles in modulating the waveforms of sperm flagella. However, a Ca2+‐binding protein in sperm flagella that regulates axonemal dyneins has not been fully characterized. Results. We identified a novel neuronal calcium sensor family protein, named calaxin (Ca2+‐binding axonemal protein), in sperm flagella of the ascidian Ciona intestinalis. Calaxin has three EF‐hand Ca2+‐binding motifs, and its orthologues are present in metazoan species, but not in yeast, green algae or plant. Immunolocalization revealed that calaxin is localized near the outer arm of the sperm flagellar axonemes. Moreover, it is distributed in adult tissues bearing epithelial cilia. An in vitro binding experiment indicated that calaxin binds to outer arm dynein. A cross‐linking experiment showed that calaxin binds to β‐tubulin in situ. Overlay experiments further indicated that calaxin binds the β‐dynein heavy chain of outer arm dynein in the presence of Ca2+. Conclusions. These results suggest that calaxin is a potential Ca2+‐dependent modulator of outer arm dynein in metazoan cilia and flagella. 相似文献
125.
126.
Marian Kruszynski Nicole Stowell Anuk Das Jonathan Seideman Ping Tsui Michael Brigham-Burke Jennifer F Nemeth Raymond Sweet George A Heavner 《Journal of peptide science》2006,12(1):25-32
Novel analogs of human monocyte chemoattractant protein 1 (MCP-1) were designed, synthesized and characterized to be used as tools to generate monoclonal antibodies as potential human therapeutics. MCP-1 and three analogs were synthesized by step-wise Fmoc solid phase synthesis. After oxidation to form the two-disulfide bonds, affinity chromatography using an immobilized mouse anti-human MCP-1 monoclonal antibody (mAb) was utilized for a simple and highly effective purification procedure for the proteins. The final products were extensively characterized and compared with recombinant human MCP-1 (rhMCP-1). All proteins showed identical binding with mouse anti-human MCP-1 mAbs as measured by surface plasmon resonance. Synthetic MCP-1 and the analogs were comparable to recombinant MCP-1 in competition radio-ligand binding to CCR2 receptors on THP-1 cells, and MCP-1-induced, calcium mobilization and chemotaxis assays. 相似文献
127.
Mira Stone Olson Roseanne M. Ford James A. Smith Erik J. Fernandez 《Bioremediation Journal》2006,10(1-2):13-23
The impact of bacterial chemotaxis on in situ ground-water bioremediation remains an unanswered question. Although bacteria respond to chemical gradients in aqueous environments and under no-flow conditions, it is unclear whether they can also respond in porous media with advective flow to improve overall contaminant degradation. The effect of chemotaxis is most profound in regions with sharp chemical gradients, most notably around residual nonaqueous phase liquid (NAPL) ganglia and surrounding clay lenses or aquitards with trapped contamination. The purpose of this study is to simulate bacterial transport through a two-dimensional subsurface environment, containing one region of low permeability with trapped contaminant surrounded above and below by two regions of higher permeability. Using mathematical predictions of the effect of pore size on measured bacterial transport parameters, the authors observe a 50% decrease in both motility and chemotaxis in the finer-grained, low-permeability porous medium. The authors simulate how chemotaxis affects bacterial migration to the contaminated region under various flow and initial conditions. Results indicate that bacteria traveling through a high-permeability region with advective flow can successfully migrate toward and accumulate around a contaminant diffusing from a lower permeability region. 相似文献
128.
Myosin II plays critical roles in events such as cytokinesis, chemotactic migration, and morphological changes during multicellular development. The amoeba Dictyostelium discoideum provides a simple system for the study of this contractile protein. In this system, myosin II filament assembly is regulated by myosin heavy chain (MHC) phosphorylation in the tail region of the molecule. Earlier studies identified an alpha-kinase, MHC kinase A (MHCK A), which phosphorylates three mapped threonine residues in the myosin tail, driving myosin disassembly. Using molecular and genomic approaches, we have identified a series of related kinases in Dictyostelium. The enzyme MHCK B shares with MHCK A a domain organization that includes a highly novel catalytic domain coupled to a carboxyl-terminal WD repeat domain. We have engineered, expressed, and purified a FLAG-tagged version of the novel kinase. In the present study, we report detailed biochemical and cellular studies documenting that MHCK B plays a physiological role in the control of Dictyostelium myosin II assembly and disassembly during the vegetative life of Dictyostelium amoebae. The presented data supports a model of multiple related MHCKs in this system, with different regulatory mechanisms and pathways controlling each enzyme. 相似文献
129.
Caenorhabditis elegans is a model organism in biology, yet despite the tremendous information generated from genetic, genomic and functional analyses, C. elegans has rarely been used to address questions in ecological genetics. Here, we analyse genetic variation for chemosensory behaviour, an ecologically important trait that is also genetically well characterized, at both the phenotypic and molecular levels within three species of the genus Caenorhabditis. We show that the G-protein ODR-3 plays an important role in chemosensory avoidance behaviour and identify orthologues of odr-3 in C. briggsae and C. remanei. Both quantitative genetic analysis of chemosensory behaviour and molecular population genetic analysis of odr-3 show that there is little genetic variation among a worldwide collection of isolates of the primarily selfing C. elegans, whereas there is substantially more variation within a single population of the outcrossing C. remanei. Although there are a large number of substitutions at silent sites within odr-3 among the three species, molecular evolution at the protein level is extremely conserved, suggesting that odr-3 plays an important role in cell signalling during chemosensation and/or neuronal cilia development in C. remanei and in C. briggsae as it does in C. elegans. Our results suggest that C. remanei may be a more suitable subject for ecological and evolutionary genetic studies than C. elegans. 相似文献
130.
Recently, we cloned and sequenced the cDNA of allurin, a sperm chemoattractant isolated from the jelly of Xenopus laevis eggs [Proc. Natl. Acad. Sci. U.S.A. 78 (2001) 11205]. In this report, we demonstrate that allurin mRNA is expressed almost exclusively in the oviduct and that its expression is increased 2.5-fold by human chorionic gonadotropin over a 12-h period. Both dot blots and immunocytochemistry show that allurin is secreted from the upper two thirds of the oviduct that includes the pars recta and the proximal pars convoluta. Allurin appears to be deposited on the ciliated surfaces of luminal epithelial cells that come in direct contact with eggs as they move through the oviduct. Immune staining also demonstrates the presence of allurin in the serosal capsule of the oviduct. In contrast, allurin is not found within the tubular jelly-secreting glands or ducts that constitute a major portion of the oviduct wall. Therefore, we hypothesize that allurin is synthesized by nonciliated secretory cells in the luminal epithelium of the oviduct, is displayed on the ciliary layer and then mechanically mixed with jelly, and applied to eggs as they progress down the oviduct. This hypothesis is consistent with the fact that eggs progressing down the oviduct initially show evidence of allurin being incorporated into the J1 layer. Subsequently, allurin within J1 diffuses outward to J3 and eggs stored in the uterus now demonstrate a J3 localization of this chemoattractant. 相似文献