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81.
82.
Limnology - In the original publication of the article the sentence under the heading “Primer and probe design” was published incorrectly. The correct sentence is given in this correction. 相似文献
83.
Seven ascidiacyclamide [cyclo(–Ile–oxazoline–d ‐Val–thiazole–)2] (ASC) analogues incorporating the β‐amino acids βIle, βoxazoline, and/or d ‐βVal were synthesized. We then investigated the effects of the position and number of incorporated β‐amino acids on the structure, cytotoxicity, and copper binding by these seven analogues. The structural analyses revealed that both βIle and d ‐βVal favor a gauche‐type θ torsion angles, while βoxazoline favors a trans‐type θ torsion angle. Expansion of the macrocycle by incorporation of βIle or d ‐βVal readily induced molecular folding. On the other hand, the incorporation of two βoxazoline residues strongly extended the peptide conformation, and the incorporation of one was sufficient for the moderate restriction important for conformational equilibrium and cytotoxicity. Despite expansion of the macrocycles, the structure‐cytotoxicity relationships were largely maintained. In studies of complexation of the analogues with Cu (II) ion, the position and number of incorporated β‐amino acids had a large impact on the structure of the metal complex and may contribute to its stabilization. 相似文献
84.
Toshio Naka Takashi Doi Takeshi Tsujino Tohru Masuyama Seinosuke Kawashima 《Preparative biochemistry & biotechnology》2013,43(2):87-99
Abstract Viral vector systems are efficient for transfection of foreign genes into many tissues. Especially, retrovirus based vectors integrate the transgene into the genome of the target cells, which can sustain long term expression. However, it has been demonstrated that the transduction efficiency using retrovirus is relatively lower than those of other viruses. Ultrasound was recently reported to increase gene expression using plasmid DNA, with or without, a delivery vehicle. However, there are no reports, which show an ultrasound effect to retrovirus‐mediated gene transfer efficiency. Retrovirus‐mediated gene transfer systems were used for transfection of 293T cells, bovine aortic endothelial cells (BAECs), rat aortic smooth muscle cells (RASMCs), and rat skeletal muscle myoblasts (L6 cells) with β‐galactosidase (β‐Gal) genes. Transduction efficiency and cell viability assay were performed on 293T cells that were exposed to varying durations (5 to 30 seconds) and power levels (1.0 watts/cm2 to 4.0 watts/cm2) of ultrasound after being transduced by a retrovirus. Effects of ultrasound to the retrovirus itself was evaluated by transduction efficiency of 293T cells. After exposure to varying power levels of ultrasound to a retrovirus for 5 seconds, 293T cells were transduced by a retrovirus, and transduction efficiency was evaluated. Below 1.0 watts/cm2 and 5 seconds exposure, ultrasound showed increased transduction efficiency and no cytotoxicity to 293T cells transduced by a retrovirus. Also, ultrasound showed no toxicity to the virus itself at the same condition. Exposure of 5 seconds at the power of 1.0 watts/cm2 of an ultrasound resulted in significant increases in retrovirus‐mediated gene expression in all four cell types tested in this experiment. Transduction efficiencies by ultrasound were enhanced 6.6‐fold, 4.8‐fold, 2.3‐fold, and 3.2‐fold in 293T cells, BAECs, RASMCs, and L6 cells, respectively. Furthermore, β‐Gal activities were also increased by the retrovirus with ultrasound exposure in these cells. Adjunctive ultrasound exposure was associated with enhanced retrovirus‐mediated transgene expression in vitro. Ultrasound associated local gene therapy has potential for not only plasmid‐DNA‐, but also retrovirus‐mediated gene transfer. 相似文献
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Masako Nomaguchi Masaru Yokoyama Ken Kono Emi E. Nakayama Tatsuo Shioda Naoya Doi Sachi Fujiwara Akatsuki Saito Hirofumi Akari Kei Miyakawa Akihide Ryo Hirotaka Ode Yasumasa Iwatani Tomoyuki Miura Tatsuhiko Igarashi Hironori Sato Akio Adachi 《Journal of virology》2013,87(21):11447-11461
Human immunodeficiency virus type 1 (HIV-1) replication in macaque cells is restricted mainly by antiviral cellular APOBEC3, TRIM5α/TRIM5CypA, and tetherin proteins. For basic and clinical HIV-1/AIDS studies, efforts to construct macaque-tropic HIV-1 (HIV-1mt) have been made by us and others. Although rhesus macaques are commonly and successfully used as infection models, no HIV-1 derivatives suitable for in vivo rhesus research are available to date. In this study, to obtain novel HIV-1mt clones that are resistant to major restriction factors, we altered Gag and Vpu of our best HIV-1mt clone described previously. First, by sequence- and structure-guided mutagenesis, three amino acid residues in Gag-capsid (CA) (M94L/R98S/G114Q) were found to be responsible for viral growth enhancement in a macaque cell line. Results of in vitro TRIM5α susceptibility testing of HIV-1mt carrying these substitutions correlated well with the increased viral replication potential in macaque peripheral blood mononuclear cells (PBMCs) with different TRIM5 alleles, suggesting that the three amino acids in HIV-1mt CA are involved in the interaction with TRIM5α. Second, we replaced the transmembrane domain of Vpu of this clone with the corresponding region of simian immunodeficiency virus SIVgsn166 Vpu. The resultant clone, MN4/LSDQgtu, was able to antagonize macaque but not human tetherin, and its Vpu effectively functioned during viral replication in a macaque cell line. Notably, MN4/LSDQgtu grew comparably to SIVmac239 and much better than any of our other HIV-1mt clones in rhesus macaque PBMCs. In sum, MN4/LSDQgtu is the first HIV-1 derivative that exhibits resistance to the major restriction factors in rhesus macaque cells. 相似文献
87.
Taro Saito Masashi Yano Yusei Kawai Akiko Asada Mitsuhito Wada Hirofumi Doi Shin-ichi Hisanaga 《The Journal of biological chemistry》2013,288(45):32433-32439
Cyclin-dependent kinase 5 (Cdk5) is a brain-specific membrane-bound protein kinase that is activated by binding to the p35 or p39 activator. Previous studies have focused on p35-Cdk5, and little is known regarding p39-Cdk5. The lack of functional understanding of p39-Cdk5 is due, in part, to the labile property of p39-Cdk5, which dissociates and loses kinase activity in nonionic detergent conditions. Here we investigated the structural basis for the instability of p39-Cdk5. p39 and p35 contain N-terminal p10 regions and C-terminal Cdk5 activation domains (AD). Although p35 and p39 show higher homology in the C-terminal AD than the N-terminal region, the difference in stability is derived from the C-terminal AD. Based on the crystal structures of the p25 (p35 C-terminal region including AD)-Cdk5 complex, we simulated the three-dimensional structure of the p39 AD-Cdk5 complex and found differences in the hydrogen bond network between Cdk5 and its activators. Three amino acids of p35, Asp-259, Asn-266, and Ser-270, which are involved in hydrogen bond formation with Cdk5, are changed to Gln, Gln, and Pro in p39. Because these three amino acids in p39 do not participate in hydrogen bond formation, we predicted that the number of hydrogen bonds between p39 and Cdk5 was reduced compared with p35 and Cdk5. Using substitution mutants, we experimentally validated that the difference in the hydrogen bond network contributes to the different properties between Cdk5 and its activators. 相似文献
88.
89.
Two kinds of carboxypeptidases (F–I, F–II) were purified from the sarcocarp of watermelon (Citrullus vulgaris, var. Shimao). F–I was not purified to homogeneity. F–II was homogeneous on ultracentrifugal analysis, but a trace of impurity was detected at high concentrations by disc electrophoresis.F–I was optimally active and stable at pH 5.0~5.5 and was strongly inhibited by DFP and HgCl2, but not by EDTA. The molecular weight and isoelectric point were 89,000 and 4.4, respectively.F–II was optimally active at pH 5.0 ~ 5.5 and was most stable at pH 5.5 ~ 7.0. It was completely inhibited by DFP and HgCl2, but not by EDTA and 1, 10-phenanthroline, and it hydrolyzed an oligopeptide containing proline, glutamic acid, lysine and several neutral amino acids, sequentially from the C-terminal. The molecular weight and isolelectric point were 110,000 (5.1 S) and 5.0, respectively.The similarity of enzymatic properties of both the present enzymes to those of other plant carboxypeptidases and pig kidney cathepsin A are discussed. 相似文献
90.