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991.
Summary Proteasomes, also known as multicatalytic proteinase complexes, were localized in suspension cells of potato (Solanum tuberosum) by direct immunofluorescence using polyclonal antibodies labelled with fluorescein isothiocyanate. The method used allows an estimate of relative amounts of proteasomal antigens in different cell components. Proteasomes are present in the nuclei and the cytoplasm. The nucleoplasm contains small areas of weak fluorescence. The peripheral cytoplasm and possibly elements of the cytoskeleton show higher fluorescence than other parts of the cytoplasm. This indicates a localization of proteasomes similar to that known from animal cells.Abbreviations DMSO
dimethylsulfoxide
- EGTA
ethyleneglycol-bis-(-aminoethylether)-N,N,N,N-tetra acetic acid
- FITC
fluorescein isothiocyanate
- PBS
phosphate buffered saline
- PIPES
piperazine-1,4-bis-(2-ethanesulfonic acid) 相似文献
992.
Significant hydrogen exchange protection in GroEL-bound DHFR is maintained during iterative rounds of substrate cycling. 下载免费PDF全文
M. Gross C. V. Robinson M. Mayhew F. U. Hartl S. E. Radford 《Protein science : a publication of the Protein Society》1996,5(12):2506-2513
An unresolved key issue in the mechanism of protein folding assisted by the molecular chaperone GroEL is the nature of the substrate protein bound to the chaperonin at different stages of its reaction cycle. Here we describe the conformational properties of human dihydrofolate reductase (DHFR) bound to GroEL at different stages of its ATP-driven folding reaction, determined by hydrogen exchange labeling and electrospray ionization mass spectrometry. Considerable protection involving about 20 hydrogens is observed in DHFR bound to GroEL in the absence of ATP. Analysis of the line width of peaks in the mass spectra, together with fluorescence quenching and ANS binding studies, suggest that the bound DHFR is partially folded, but contains stable structure in a small region of the polypeptide chain. DHFR rebound to GroEL 3 min after initiating its folding by the addition of MgATP was also examined by hydrogen exchange, fluorescence quenching, and ANS binding. The results indicate that the extent of protection of the substrate protein rebound to GroEL is indistinguishable from that of the initial bound state. Despite this, small differences in the quenching coefficient and ANS binding properties are observed in the rebound state. On the basis of these results, we suggest that GroEL-assisted folding of DHFR occurs by minor structural adjustments to the partially folded substrate protein during iterative cycling, rather than by complete unfolding of this protein substrate on the chaperonin surface. 相似文献
993.
994.
Cryptosporidium spp. in domestic dogs: the "dog" genotype 总被引:3,自引:0,他引:3
Morgan UM Xiao L Monis P Fall A Irwin PJ Fayer R Denholm KM Limor J Lal A Thompson RC 《Applied and environmental microbiology》2000,66(5):2220-2223
Genetic and phylogenetic characterization of Cryptosporidium isolates at two loci (18S rRNA gene and heat shock gene) from both Australian and United States dogs demonstrated that dog-derived Cryptosporidium isolates had a distinct genotype which is conserved across geographic areas. Phylogenetic analysis provided support for the idea that the "dog" genotype is, in fact, a valid species. 相似文献
995.
Phylogenetic relationships of Cryptosporidium parasites based on the 70-kilodalton heat shock protein (HSP70) gene 总被引:10,自引:0,他引:10
Sulaiman IM Morgan UM Thompson RC Lal AA Xiao L 《Applied and environmental microbiology》2000,66(6):2385-2391
We have characterized the nucleotide sequences of the 70-kDa heat shock protein (HSP70) genes of Cryptosporidium baileyi, C. felis, C. meleagridis, C. muris, C. serpentis, C. wrairi, and C. parvum from various animals. Results of the phylogenetic analysis revealed the presence of several genetically distinct species in the genus Cryptosporidium and eight distinct genotypes within the species C. parvum. Some of the latter may represent cryptic species. The phylogenetic tree constructed from these sequences is in agreement with our previous results based on the small-subunit rRNA genes of Cryptosporidium parasites. The Cryptosporidium species formed two major clades: isolates of C. muris and C. serpentis formed the first major group, while isolates of C. felis, C. meleagridis, C. wrairi, and eight genotypes of C. parvum formed the second major group. Sequence variations were also observed between C. muris isolates from ruminants and rodents. The HSP70 gene provides another useful locus for phylogenetic analysis of the genus Cryptosporidium. 相似文献
996.
Norlin M Andersson U Björkhem I Wikvall K 《The Journal of biological chemistry》2000,275(44):34046-34053
A 7 alpha-hydroxylation is necessary for conversion of both cholesterol and 27-hydroxycholesterol into bile acids. According to current theories, cholesterol 7 alpha-hydroxylase (CYP7A) is responsible for the former and oxysterol 7 alpha-hydroxylase (CYP7B) for the latter reaction. CYP7A is believed to have a very high substrate specificity whereas CYP7B is active toward oxysterols, dehydroepiandrosterone, and pregnenolone. In the present study, 7 alpha-hydroxylation of various oxysterols in liver and kidney was investigated. Surprisingly, human cholesterol 7 alpha-hydroxylase, CYP7A, expressed as a recombinant in Escherichia coli and COS cells, was active toward 20(S)-hydroxycholesterol, 25-hydroxycholesterol, and 27-hydroxycholesterol. This enzyme has previously been thought to be specific for cholesterol and cholestanol. A partially purified and reconstituted cholesterol 7 alpha-hydroxylase enzyme fraction from pig liver showed 7 alpha-hydroxylase activity toward the same oxysterols as metabolized by expressed recombinant human and rat CYP7A. The 7 alpha-hydroxylase activity toward 20(S)-hydroxycholesterol, 25-hydroxycholesterol, and 27-hydroxycholesterol in rat liver was significantly increased by treatment with cholestyramine, an inducer of CYP7A. From the present results it may be concluded that CYP7A is able to function as an oxysterol 7 alpha-hydroxylase, in addition to the previously known human oxysterol 7 alpha-hydroxylase, CYP7B. These findings may have implications for oxysterol-mediated regulation of gene expression and for pathways of bile acid biosynthesis. A possible use of 20(S)-hydroxycholesterol as a marker substrate for CYP7A is proposed. 相似文献
997.
998.
Direct action of endocrine disrupting chemicals on human sperm 总被引:1,自引:0,他引:1
Dorte L Egeberg Luis Alvarez Christoph Brenker Anders Rehfeld Hanne Frederiksen Benjamin Wäschle U Benjamin Kaupp Melanie Balbach Dagmar Wachten Niels E Skakkebaek Kristian Almstrup Timo Strünker 《EMBO reports》2014,15(7):758-765
Synthetic endocrine disrupting chemicals (EDCs), omnipresent in food, household, and personal care products, have been implicated in adverse trends in human reproduction, including infertility and increasing demand for assisted reproduction. Here, we study the action of 96 ubiquitous EDCs on human sperm. We show that structurally diverse EDCs activate the sperm‐specific CatSper channel and, thereby, evoke an intracellular Ca2+ increase, a motility response, and acrosomal exocytosis. Moreover, EDCs desensitize sperm for physiological CatSper ligands and cooperate in low‐dose mixtures to elevate Ca2+ levels in sperm. We conclude that EDCs interfere with various sperm functions and, thereby, might impair human fertilization. 相似文献
999.
Novel VEGF family members: VEGF-B, VEGF-C and VEGF-D 总被引:6,自引:0,他引:6
Vascular endothelial growth factors (VEGFs) constitute a group of structurally and functionally related growth factors that modulate many important physiological functions of endothelial cells. Currently, five different mammalian VEGFs have been identified and they all show unique temporal and spatial expression patterns, receptor specificity and function. The VEGFs may play pivotal roles in formation of the vascular systems during embryonic development, in regulation of capillary growth in normal and pathological conditions in adults, and in the maintenance of the normal vasculature. In the future, the VEGFs and their receptors may become important therapeutic tools in treatment of conditions characterized by aberrant or defective formation of blood vessels and lymphatic vessels. 相似文献
1000.
U Seidel E Bober B Winter S Lenz P Lohse H W Goedde K H Grzeschik H H Arnold 《Gene》1988,66(1):135-146
A set of cDNA clones coding for alkali myosin light chains (AMLC) was isolated from fetal human skeletal muscle. Nucleotide sequence analysis and RNA expression patterns of individual clones revealed related sequences corresponding to (i) fast fiber type MLC1 and MLC3; (ii) the embryonic MLC that is also expressed in fetal ventricle and adult atrium (MLCemb); and (iii) a nonsarcomeric MLC isoform that is found in all nonmuscle cell types and smooth muscle. The AMLC gene family in man comprises unique copies for MLC1, MLC3 and MLCemb, and multiple copies for the nonsarcomeric MLC genes. The gene coding for MLC1 and MLC3 is located on human chromosome 2. 相似文献