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81.
Takaaki Kameji Yasuko Murakami Kazunobu Fujita Shin-Ichi Hayashi 《Biochimica et Biophysica Acta (BBA)/General Subjects》1982,717(1):111-117
Ornithine decarboxylase (EC 4.1.1.17) was purified to near homogeniety from livers of thioacetamide- and dl-α-hydrazino-δ-aminovaleric acid-treated rats by using three types of affinity chromatography with pyridoxamine phosphate-Sepharose, pyridoxamine phosphate-dipropylenetriamine-Sepharose and heparin-Sepharose. This procedure gave a purification of about 3.5·105-fold with an 8% yield; the specific activity of the final enzyme preparation was 1,1·106 nmol CO2/h per mg protein. The purified enzyme gave a single band of protein which coincided with activity peak on polyacrylamide gel electrophoresis and also gave a single major band on SDS-polyacrylamide gel electrophoresis. A single precipitin line was formed between the purified enzyme and an antiserum raised against a partially purified enzyme, on Ouchterlony immunodiffusion. The molecular weight of the enzyme was estimated to be 105 000 by polyacrylamide gel electrophoresis at several different gel concentrations; the dissociated subunits had molecular weights of 50 000 on SDS-polyacrylmide gels. The isoelectric point of the enzyme was pH 4.1. 相似文献
82.
Tochio N Umehara T Koshiba S Inoue M Yabuki T Aoki M Seki E Watanabe S Tomo Y Hanada M Ikari M Sato M Terada T Nagase T Ohara O Shirouzu M Tanaka A Kigawa T Yokoyama S 《Structure (London, England : 1993)》2006,14(3):457-468
SWIRM is an evolutionarily conserved domain involved in several chromatin-modifying complexes. Recently, the LSD1 protein, which bears a SWIRM domain, was found to be a demethylase for Lys4-methylated histone H3. Here, we report a solution structure of the SWIRM domain of human LSD1. It forms a compact fold composed of 6 alpha helices, in which a 20 amino acid long helix (alpha4) is surrounded by 5 other short helices. The SWIRM domain structure could be divided into the N-terminal part (alpha1-alpha3) and the C-terminal part (alpha4-alpha6), which are connected to each other by a salt bridge. While the N-terminal part forms a SWIRM-specific structure, the C-terminal part adopts a helix-turn-helix (HTH)-related fold. We discuss a model in which the SWIRM domain acts as an anchor site for a histone tail. 相似文献
83.
Shintaro Kobayashi Tadaki Suzuki Manabu Igarashi Yasuko Orba Noriko Ohtake Keita Nagakawa Kenichi Niikura Takashi Kimura Harumi Kasamatsu Hirofumi Sawa 《PloS one》2013,8(10)
The capsid of the human polyomavirus JC virus (JCV) consists of 72 pentameric capsomeres of a major structural protein, Vp1. The cysteine residues of the related Vp1 of SV40 are known to contribute to Vp1 folding, pentamer formation, pentamer-pentamer contacts, and capsid stabilization. In light of the presence of a slight structural difference between JCV Vp1 and SV40 counterpart, the way the former folds could be either different from or similar to the latter. We found a difference: an important contribution of Vp1 cysteines to the formation of infectious virions, unique in JCV and absent in SV40. Having introduced amino acid substitution at each of six cysteines (C42, C80, C97, C200, C247, and C260) in JCV Vp1, we found that, when expressed in HeLa cells, the Vp1 level was decreased in C80A and C247A mutants, and remained normal in the other mutants. Additionally, the C80A and C247A Vp1-expressing cell extracts did not show the hemagglutination activity characteristic of JCV particles. The C80A and C247A mutant Vp1s were found to be less stable than the wild-type Vp1 in HeLa cells. When produced in a reconstituted in vitro protein translation system, these two mutant proteins were stable, suggesting that some cellular factors were responsible for their degradation. As determined by their sucrose gradient sedimentation profiles, in vitro translated C247A Vp1 formed pentamers, but in vitro translated C80A Vp1 was entirely monomeric. When individually incorporated into the JCV genome, the C80A and C247A mutants, but not the other Vp1 cysteine residues mutants, interfered with JCV infectivity. Furthermore, the C80A, but not the C247A, mutation prevented the nuclear localization of Vp1 in JCV genome transfected cells. These findings suggest that C80 of JCV Vp1 is required for Vp1 stability and pentamer formation, and C247 is involved in capsid assembly in the nucleus. 相似文献
84.
The dynamic behavior of organelles is essential for plant survival under various environmental conditions. Plant organelles, with various functions,migrate along actin filaments and contact other types of organelles, leading to physical interactions at a specific site called the membrane contact site. Recent studies have revealed the importance of physical interactions in maintaining efficient metabolite flow between organelles.In this review, we first summarize peroxisome function under different environmental conditions and growth stages to understand organelle interactions. We then discuss current knowledge regarding the interactions between peroxisome and other organelles, i.e., the oil bodies, chloroplast, and mitochondria from the perspective of metabolic and physiological regulation, with reference to various organelle interactions and techniques for estimating organelle interactions occurring in plant cells. 相似文献
85.
Nagoshi Y Kuwasako K Cao YN Kitamura K Eto T 《Biochemical and biophysical research communications》2004,314(4):1057-1063
We examined the effects of recombinant human C-reactive protein (rhCRP) on atherosclerosis-related factors in cultured human coronary artery endothelial and smooth muscle cells (HCAECs and HCASMCs). After removing endotoxin from commercial rhCRP preparations using the appropriate column, the purified (P)-rhCRP retained the ability to Ca(2+)-dependently bind to phosphorylcholine, but did not augment the secretion of interleukin-6 and MCP-1 from HCAECs, as non-purified (NP)-rhCRP did. By contrast, P-rhCRP elicited 2- to 3-fold increases in the secretion of both hormones from HCASMCs, though the effect was smaller than that obtained with NP-rhCRP. Production of PAI-1 and endothelin-1 was little affected by either rhCRP preparation in either cell type. In addition, P-rhCRP dose-dependently diminished adrenomedullin release from both cell types, but did not affect adrenomedullin receptor expression or function. Our findings highlight the importance of removing endotoxin from commercial rCRP preparations and show that hCRP elicits atherogenic responses from HCASMCs, but not HCAECs. 相似文献
86.
The objective of this study was to develop a method for whole-mount in situ hybridization (WISH) by using elongation factor-1 (EF-1) riboprobes in the red alga Porphyra yezoensis Ueda. Several modifications to the general WISH protocol, such as use of a short-length probe, performing partial digestion of the cell wall, optimization of proteinase K concentration and additional washing steps after hybridization were essential to reduce non-specific staining and to obtain sufficient quality of data. This protocol made it possible to detect a specific signal as a positive control in WISH assays of P. yezoensis. 相似文献
87.
True hermaphroditism with 46,X,+22p/46,XY and gonadal mosaicism detected by fluorescence in situ hybridization 总被引:1,自引:0,他引:1
A Japanese girl was diagnosed as true hermaphroditism with 46,X,+mar/46,XY and the marker chromosome was determined on the short arm of chromosome 22 without alpha-satellite by fluorescence in situ hybridization (FISH) and spectral karyotyping (SKY) methods. At birth, she showed intersexual external genitalia, urethral-vaginal fistula and right inguinal hernia. The right gonad was revealed as an ovotestis, and the left was as an undifferentiated testis. The gonadal mosaicism was demonstrated directly in gonadal tissue by interphase FISH. 相似文献
88.
Iwakiri Y Tsai MH McCabe TJ Gratton JP Fulton D Groszmann RJ Sessa WC 《American journal of physiology. Heart and circulatory physiology》2002,282(6):H2084-H2090
Akt, also known as protein kinase B, is a serine/threonine kinase. Akt becomes active when phosphorylated by the activation of receptor tyrosine kinases, G protein-coupled receptors, and mechanical forces such as shear stress. Studies in vitro have shown that Akt can directly phosphorylate endothelial nitric oxide (NO) synthase (eNOS) and activate the enzyme, leading to NO production. The aim of this study was to test the hypothesis that the phosphorylation of eNOS plays a role in the enhanced NO production observed in early portal hypertension. Male Sprague-Dawley rats were subjected to either sham or portal vein ligation (PVL), and mesenteric arterial beds were used for ex vivo perfusion studies. Mesenteric arterial beds from PVL rats had an approximately 60-70% decrease in response to methoxamine (an alpha(1)-agonist and vasoconstrictor) compared with the sham group (P < 0.01). When N(G)-monomethyl-L-arginine (a NOS inhibitor) was added to the perfusion, the difference in perfusion pressure between the two groups was abolished, suggesting that enhanced NO production in the PVL group blunted the response to the vasoconstrictor. The reduced responsiveness in PVL was not due to changes in eNOS expression but was due to an increase in enzyme-specific activity, suggesting posttranslational modification of eNOS. The phosphorylation of eNOS at Ser(1176) was significantly increased by twofold (P < 0.05) in the PVL group. Furthermore, PVL significantly increased Akt phosphorylation (an active form of Akt) by threefold (P < 0.05). When vessels were treated with wortmannin (10 nM) to block the phosphatidylinositol-3-OH-kinase/Akt pathway, NO-induced vasodilatation was significantly reduced. These results suggest that the phosphorylation of eNOS by Akt activates the enzyme and may be the first step leading to an initial increase in NO production in portal hypertension. 相似文献
89.
Isolation and identification of EG-VEGF/prokineticins as cognate ligands for two orphan G-protein-coupled receptors 总被引:8,自引:0,他引:8
Masuda Y Takatsu Y Terao Y Kumano S Ishibashi Y Suenaga M Abe M Fukusumi S Watanabe T Shintani Y Yamada T Hinuma S Inatomi N Ohtaki T Onda H Fujino M 《Biochemical and biophysical research communications》2002,293(1):396-402
Endocrine gland-derived vascular endothelial growth factor (EG-VEGF, identical to prokineticin 1) is a novel peptide recently identified as a selective mitogen for endocrine gland endothelial cells. The present study demonstrates that EG-VEGF/prokineticin 1 and a peptide closely related to EG-VEGF, prokineticin 2, are cognate ligands of two orphan G-protein-coupled receptors designated ZAQ (=EG-VEGF/PK-R1) and I5E (=EG-VEGF/PK-R2). EG-VEGF/prokineticin 1 and prokineticin 2 induced a transient increase in intracellular calcium ion concentration ([Ca(2+)](i)) with nanomolar potency in Chinese hamster ovary (CHO) cells expressing EG-VEGF/PK-R1 and -R2 and bind to these cells with high affinity and with different receptor selectivity. EG-VEGF/prokineticins provoke rapid phosphorylation of p44/42 MAP kinase and DNA synthesis in the bovine adrenal capillary endothelial cells (BACE). The mRNAs of both EG-VEGF/PK-R1 and -R2 were expressed in BACE. The identification of the receptors for EG-VEGF/prokineticins may provide a novel molecular basis for the regulation of angiogenesis in endocrine glands. 相似文献
90.
KIFC3, a microtubule (MT) minus end-directed kinesin superfamily protein, is expressed abundantly and is associated with the Golgi apparatus in adrenocortical cells. We report here that disruption of the kifC3 gene induced fragmentation of the Golgi apparatus when cholesterol was depleted. Analysis of the reassembly process of the Golgi apparatus revealed bidirectional movement of the Golgi fragments in both wild-type and kifC3-/- cells. However, we observed a markedly reduced inwardly directed motility of the Golgi fragments in cholesterol-depleted kifC3-/- cells compared with either cholesterol-depleted wild-type cells or cholesterol-replenished kifC3-/- cells. These results suggest that (a) under the cholesterol-depleted condition, reduced inwardly directed motility of the Golgi apparatus results in the observed Golgi scattering phenotype in kifC3-/- cells, and (b) cholesterol is necessary for the Golgi fragments to attain sufficient inwardly directed motility by MT minus end-directed motors other than KIFC3, such as dynein, in kifC3-/- cells. Furthermore, we showed that Golgi scattering was much more drastic in kifC3-/- cells than in wild-type cells to the exogenous dynamitin expression even in the presence of cholesterol. These results collectively demonstrate that KIFC3 plays a complementary role in Golgi positioning and integration with cytoplasmic dynein. 相似文献