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91.
Takata T Kimura Y Ohnuma Y Kawawaki J Kakiyama Y Tanaka K Kakizuka A 《Journal of structural biology》2012,179(2):93-103
VCP/p97/Cdc48 is a hexameric ring-shaped AAA ATPase that participates in a wide variety of cellular functions. VCP is a very abundant protein in essentially all types of cells and is highly conserved among eukaryotes. To date, 19 different single amino acid-substitutions in VCP have been reported to cause IBMPFD (inclusion body myopathy associated with Paget disease of bone and frontotemporal dementia), an autosomal dominant inherited human disease. Moreover, several similar single amino acid substitutions have been proposed to associate with a rare subclass of familial ALS. The mechanisms by which these mutations contribute to the pathogenesis are unclear. To elucidate potential functional differences between wild-type and pathogenic VCPs, we expressed both VCPs in yeast cdc48 mutants. We observed that all tested pathogenic VCPs suppressed the temperature-sensitive phenotype of cdc48 mutants more efficiently than wild-type VCP. In addition, pathogenic VCPs, but not wild-type VCP, were able to rescue a lethal cdc48 disruption. In yeast, pathogenic VCPs, but not wild-type VCP, formed apparent cytoplasmic foci, and these foci were transported to budding sites by the Myo2/actin-mediated transport machinery. The foci formation of pathogenic VCPs appeared to be associated with their suppression of the temperature-sensitive phenotype of cdc48 mutants. These results support the idea that the pathogenic VCP mutations create dominant gain-of-functions rather than a simple loss of functional VCP. Their unique properties in yeast could provide a convenient drug-screening system for the treatment of these diseases. 相似文献
92.
Nakamichi Y Maruyama Y Mikami B Hashimoto W Murata K 《The Journal of biological chemistry》2011,286(8):6262-6271
Pathogenic Streptococcus agalactiae produces polysaccharide lyases and unsaturated glucuronyl hydrolase (UGL), which are prerequisite for complete degradation of mammalian extracellular matrices, including glycosaminoglycans such as chondroitin and hyaluronan. Unlike the Bacillus enzyme, streptococcal UGLs prefer sulfated glycosaminoglycans. Here, we show the loop flexibility for substrate binding and structural determinants for recognition of glycosaminoglycan sulfate groups in S. agalactiae UGL (SagUGL). UGL also degraded unsaturated heparin disaccharides; this indicates that the enzyme released unsaturated iduronic and glucuronic acids from substrates. We determined the crystal structures of SagUGL wild-type enzyme and both substrate-free and substrate-bound D175N mutants by x-ray crystallography and noted that the loop over the active cleft exhibits flexible motion for substrate binding. Several residues in the active cleft bind to the substrate, unsaturated chondroitin disaccharide with a sulfate group at the C-6 position of GalNAc residue. The sulfate group is hydrogen-bonded to Ser-365 and Ser-368 and close to Lys-370. As compared with wild-type enzyme, S365H, S368G, and K370I mutants exhibited higher Michaelis constants toward the substrate. The conversion of SagUGL to Bacillus sp. GL1 UGL-like enzyme via site-directed mutagenesis demonstrated that Ser-365 and Lys-370 are essential for direct binding and for electrostatic interaction, respectively, for recognition of the sulfate group by SagUGL. Molecular conversion was also achieved in SagUGL Arg-236 with an affinity for the sulfate group at the C-4 position of the GalNAc residue. These residues binding to sulfate groups are frequently conserved in pathogenic bacterial UGLs, suggesting that the motif "R-//-SXX(S)XK" (where the hyphen and slash marks in the motif indicate the presence of over 100 residues in the enzyme and parentheses indicate that Ser-368 makes little contribution to enzyme activity) is crucial for degradation of sulfated glycosaminoglycans. 相似文献
93.
Maruyama R Takemura G Tohse N Ohkusa T Ikeda Y Tsuchiya K Minatoguchi S Matsuzaki M Fujiwara T Fujiwara H 《American journal of physiology. Heart and circulatory physiology》2006,290(4):H1493-H1502
During early apoptosis, adult cardiomyocytes show unusual beating, suggesting possible participation of abnormal Ca(2+) transients in initiation of apoptotic processes in this cell type. Simultaneously with the beating, these cells show dynamic structural alteration resulting from cytoskeletal disintegration that is quite rapid. Because of the specialized structure and extensive cytoskeleton of cardiomyocytes, we hypothesized that its degradation in so short a time would require a particularly efficient mechanism. To better understand this mechanism, we used serial video microscopy to observe beta-adrenergic stimulation-induced apoptosis in isolated adult rat cardiomyocytes while simultaneously recording intracellular Ca(2+) concentration and cell length. Trains of Ca(2+) transients and corresponding rhythmic contractions and relaxations (beating) were observed in apoptotic cells. Frequencies of Ca(2+) transients and beating gradually increased with time and were accompanied by cellular shrinkage. As the cells shrank, amplitudes of Ca(2+) transients declined and diastolic intracellular Ca(2+) concentration increased until the transients were lost. Beating and progression of apoptosis were significantly inhibited by antagonists against the L-type Ca(2+) channel (nifedipine), ryanodine receptor (ryanodine), inositol 1,4,5-trisphosphate receptor (heparin), sarco(endo)plasmic Ca(2+)-ATPase (thapsigargin), and Na(+)/Ca(2+) exchanger (KB-R7943). Electron-microscopic examination of beating cardiomyocytes revealed progressive breakdown of Z disks. Immunohistochemical analysis and Western blot confirmed that disappearance of Z disk constituent proteins (alpha-actinin, desmin, and tropomyosin) preceded degradation of other cytoskeletal proteins. It thus appears that, in adult cardiomyocyte apoptosis, Ca(2+) transients mediate apoptotic beating and efficient sarcomere destruction initiated by Z disk breakdown. 相似文献
94.
Takemura M Kazama T Sakuma K Mizushina Y Oshima T 《Bioscience, biotechnology, and biochemistry》2011,75(7):1349-1353
The DNA replication mechanism has been well established for eutherian mammals (placental mammals such as humans, mice, and cattle), but not, to date, for metatherian mammals (marsupials such as kangaroos, koalas, and opossums). In this study, we found that dehydroaltenusin, a selective inhibitor of mammalian (eutherian) DNA polymerase α, clearly suppressed the growth of metatherian (opossum and rat kangaroo) cultured cells. In cultured opossum (OK) cells, dehydroaltenusin also suppressed the progression of DNA replication. These results suggest that dehydroaltenusin inhibits metatherian as well as eutherian DNA replication. Dehydroaltenusin treatment of OK cells engendered fluctuations in the numbers of chromosomes in the OK cells as well as inhibition of cell growth and DNA replication. This suggests that partial inhibition of DNA replication by dehydroaltenusin causes chromosomal instability in cultured cells. 相似文献
95.
Koji Nagata Yukie Katayama Tomomi Sato Yeondae Kwon Takeshi Kawabata 《Journal of structural and functional genomics》2016,17(4):111-133
More than 800 G protein-coupled receptor (GPCR) genes have been discovered in the human genome. Towards the next step in GPCR research, we performed a knowledge-driven analysis of orphan class-A GPCRs that may serve as novel targets in drug discovery. We examined the relationship between 61 orphan class-A GPCR genes and diseases using the Online Mendelian Inheritance in Man (OMIM) database and the DDSS tool. The OMIM database contains data on disease-related variants of the genes. Particularly, the variants of GPR101, GPR161, and GPR88 are related to the genetic diseases: growth hormone-secreting pituitary adenoma 2, pituitary stalk interruption syndrome (not confirmed), and childhood-onset chorea with psychomotor retardation, respectively. On the other hand, the Drug Discovery and Diagnostic Support System (DDSS) tool suggests that 48 out of the 61 orphan receptor genes are related to diseases, judging from their co-occurrences in abstracts of biomedical literature. Notably, GPR50 and GPR3 are related to as many as 25 and 24 disease-associated keywords, respectively. GPR50 is related to 17 keywords of psychiatric disorders, whereas GPR3 is related to 11 keywords of neurological disorders. The aforementioned five orphan GPCRs were characterized genetically, structurally and functionally using the structural life science data cloud VaProS, so as to evaluate their potential as next targets in drug discovery. 相似文献
96.
NRSF regulates the fetal cardiac gene program and maintains normal cardiac structure and function 总被引:1,自引:0,他引:1
97.
98.
Mutated SEA-D227A-conjugated antibodies greatly enhance antitumor activity against MUC1-expressing bile duct carcinoma 总被引:8,自引:0,他引:8
Hideaki Kodama Masanori Suzuki Yu Katayose Masao Shinoda Naoki Sakurai Shin-ichi Takemura Hiroshi Yoshida Hisaaki Saeki Masahiko Ichiyama Kohei Tsumoto Ryutaro Asano Izumi Kumagai Kohzoh Imai Yuji Hinoda Seiki Matsuno Toshio Kudo 《Cancer immunology, immunotherapy : CII》2001,50(10):539-548
For the purpose of establishing a new adoptive immunotherapy for bile duct carcinoma (BDC), we have directed our attention to superantigens (SAgs), the most potent known activators of T lymphocytes. In our previous study, staphylococcal enterotoxin A (SEA) was conjugated chemically with MUSE11 mAb, which recognizes the MUC1 cancer-associated antigen, and shown to enhance the specific cytotoxic activity of T-LAK cells against MUC1-expressing BDC cells (TFK-1) in vitro and in vivo. However, it is probable that SEA might cause side-effects because of nonspecific binding to class II positive cells. In order to overcome these, we generated mutated SEA (mSEA) by changing Asp at position 227 of native SEA to Ala, which has reduced affinity to MHC class II molecules, but retains the potential for T cell activation. When mSEA-D227A was administered to rabbits to examine effects on blood pressure, 500 times more mSEA-D227A was tolerated than native SEA. This prompted us to construct a mSEA-D227A-conjugated mAb, reactive with MUC1. It augmented the antitumor activity of T-LAK cells significantly, and furthermore, mSEA-D227A could be conjugated to two bispecific antibodies, BsAb (anti-MUC1 x anti-CD3) and BsAb (anti-MUC1 x anti-CD28), which in combination had greater enhancing effects than mSEA-D227A-conjugated anti-MUC1 mAb, and combination of unconjugated BsAbs. These findings indicate a utility of mSEA-D227A-conjugated antibodies for targeted cancer immunotherapy. 相似文献
99.
Thermally induced transition between anhydrous and hydrated forms of highly crystalline beta-chitin was studied by differential thermal calorimetry (DSC) and X-ray diffraction. DSC of wet beta-chitin in a sealed pan gave two well-defined endothermic peaks at 85.2 and 104.7 degrees C on heating and one broad exothermic peak at between 60 and 0 degrees C on cooling. These peaks were highly reproducible and became more distinct after repeated heating-cooling cycles. The X-ray diffraction pattern of wet beta-chitin at elevated temperature showed corresponding changes in d-spacing between the sheets formed by stacking of chitin molecules. These phenomena clearly show that water is reversibly incorporated into the beta-chitin crystal and that the temperature change induces transitions between anhydrous, monohydrate, and dihydrate forms. The DSC behavior in heating-cooling cycles, including reversion between the two endothermic peaks, indicated that the transition between monohydrate and dihydrate was a fast and narrow-temperature process, whereas the one between the anhydrous and the monohydrate form was a slow and wide-temperature process. 相似文献
100.
Kenji Oda Katsuyuki Yamato Eiji Ohta Yasukazu Nakamura Miho Takemura Naoko Nozato Kinya Akashi Takeshi Kanegae Yutaka Ogura Takayuki Kohchi Kanji Ohyama 《Plant Molecular Biology Reporter》1992,10(2):105-163
Libraries of cosmid and plasmid clones covering the entire region of mtDNA from the liverwortMarchantia polymorpha were constructed. These clones were used for the determination of the complete nucleotide sequence of the liverwort mtDNA
totally 186,608 bp (GenBank no. M68929) and including genes for 3 species of ribosomal RNAs, 29 genes for 27 species of transfer
RNAs, and 30 genes for functionally known proteins (16 ribosomal proteins, 3 subunits of cytochromec oxidase, apocytochromeb protein, 3 subunits of H+-ATPase, and 7 subunits of NADH ubiquinone oxidoreductase). The genome also contains 32 unidentified open reading frames.
Thus the complete nucleotide sequences from both chloroplast and mitochondrial genomes have been determined in the same organism.
Plasmid clones are available upon the request.
Gene names are represented according to Lonsdale and Leaver (1988) with modifications recommended by Lonsdale (personal communication). 相似文献