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131.
132.
Glomerular endothelial cell injury plays an important role in the development and progression of diabetic nephropathy (DN). The expression and function of klotho in glomerular endothelial cells remain unclear. Thus, this study aimed to investigate the expression and the functional role of klotho in DN progression in mice and in high glucose (HG)-induced cell injury of human renal glomerular endothelial cells (HRGECs) and the underlying mechanism. In this study, HRGECs were cultured with media containing HG to induce endothelial cell injury and db/db mice were used as DN model mice. Klotho was overexpressed or knocked down in HRECs to evaluate its role in HG-induced HRGECs injury. klotho-overexpressing adenovirus (rAAV-klotho) was injected into db/db mice via the tail vein to further validate the protective effect of klotho in DN. Decreased klotho expression was observed in DN patients, DN mice, and HG-exposed HRGECs. Furthermore, klotho overexpression significantly abolished the HG-induced HRGECs injury and activation of Wnt/β-catenin pathway and RAAS. In contrast, klotho knockdown exerted the opposite effects. Moreover, klotho attenuated diabetic nephropathy in db/db mice, which was also associated with inhibition of the Wnt/β-catenin pathway and RAAS. In conclusion, klotho attenuates DN in db/db mice and ameliorates HG-induced injury of HRGECs. 相似文献
133.
【目的】为了鉴定植原体tRNA异戊烯基焦磷酸转移酶基因(tRNA-ipt)的表达及蛋白功能,探索植原体致病机理。【方法】对泡桐丛枝、桑萎缩、长春花绿变及苦楝丛枝植原体tRNA-ipt基因完整序列进行PCR扩增和生物信息学分析。对泡桐丛枝植原体tRNA-ipt基因进行原核表达并制备抗体。利用Western blot和FITC间接免疫荧光显微镜检测其在植原体中的表达。使用分光光度计分析该基因对大肠杆菌生长的影响,用ELISA测定转化菌株细胞分裂素含量。【结果】首次发现泡桐丛枝、桑萎缩、长春花绿变及苦楝丛枝植原体中完整tRNA-ipt基因,大小为876 bp,编码291个氨基酸,且N端均含有ATP/GTP结合位点保守序列(GPTASGKT)。4种植原体tRNA-IPT之间的氨基酸序列相似率为99.1%-99.5%,与同组植原体同源性在95.4%-99.3%,与其他组植原体同源性低于70%。SDS-PAGE结果显示tRNA-IPT蛋白在大肠杆菌中得到表达。首次获得泡桐丛枝植原体tRNA-IPT抗体并检测到该蛋白在泡桐发病组织中的特异表达。经过对转化菌株生长曲线及玉米素含量的测定,发现该基因能促进大肠杆菌后期生长和玉米素核苷的积累。【结论】4种植原体tRNA-ipt基因编码相同特性的功能蛋白,泡桐丛枝植原体tRNA-IPT蛋白能够在植原体中表达,根据该基因对异源菌株生长速率和激素合成的影响推断该蛋白可能参与植原体的细胞分裂素合成,在致病过程中起到重要作用。 相似文献
134.
两株淡水微囊藻的藻蓝蛋白基因间隔序列(PC-IGS)分析 总被引:2,自引:1,他引:2
对2株编号为003和004的淡水水华微囊藻(Microcystis.sp)的藻蓝蛋白基因间隔序列进行测定,获得长度均为608bp的2条序列。同时从GenBank中获取铜绿微囊藻(Microcystis aeruginosa Kütz,NCBI序列号AJ003179)及惠氏微囊藻(Microcystis wesenbergii,NCBI序列号AF385391)的序列。分别运用MEGA3及ClustalX(Version1.83)软件对这4株藻的PC-IGS序列进行碱基组成分析和序列比对。碱基组成的比对结果表明4株藻的G+C含量分别为003(50.5%),004(51.7%),铜绿微囊藻(50.7%),惠氏微囊藻(52.3%),相差范围在0.2%~1.8%之间,其结果不足以区分这四株微囊藻;序列比对则表明003号藻株与铜绿微囊藻和惠氏微囊藻的序列相似性分别为100%和88.35%,而004号藻株与铜绿微囊藻和惠氏微囊藻的序列相似性比较结果为95.13%和89.04%。此外,文章还探讨了PC-IGS序列作为微囊藻种间鉴定分子标记的可行性。 相似文献
135.
Ren X Zhao L Sivashanmugam A Miao Y Korando L Yang Z Reardon CA Getz GS Brouillette CG Jerome WG Wang J 《Biochemistry》2005,44(45):14907-14919
Apolipoprotein AI (apoAI), the major protein component of HDL, is one of the best predictors of coronary artery disease (CAD), with high apoAI and HDL levels being correlated with low occurrences of CAD. The primary function of apoAI is to recruit phospholipid and cholesterol for assembly of HDL particles. Like other exchangeable apolipoproteins, lipid-free apoAI forms a mixture of different oligomers even at 1.0 mg/mL. This self-association property of the exchangeable apolipoproteins is closely associated with the lipoprotein-binding activity of this protein family. It is unclear if the self-association property of apolipoprotein is required for its lipoprotein-binding activity. We developed a novel method for engineering an oligomeric protein to a monomeric, biologically active protein. Using this method, we generated a monomeric mouse apoAI mutant that is active. This mutant contains the first 216 residues of mouse apoAI and replaces six hydrophobic residues with either polar or smaller hydrophobic residues at the defined positions (V118A/A119S/L121Q/T191S/T195S/T199S). Cross-linking results show that this mutant is greater than 90% monomeric at 8 mg/mL. CD, DSC, and NMR results indicate that the mutant maintains an identical secondary, tertiary structure and stability as those of the wild-type mouse apoAI. Lipid-binding assays suggest that the mutant shares an equal lipoprotein-binding activity as that of the wild-type apoAI. In addition, both the monomeric mutant and the wild-type protein make nearly identical rHDL particles. With this monomeric mouse apoAI, high-quality NMR data has been collected, allowing for the NMR structural determination of lipid-free apoAI. On the basis of these results, we conclude that this apoAI mutant is a monomeric, active apoAI useful for structural determination. 相似文献
136.
Secretion, purification, and characterization of a recombinant Aspergillus oryzae tannase in Pichia pastoris 总被引:6,自引:0,他引:6
Zhong X Peng L Zheng S Sun Z Ren Y Dong M Xu A 《Protein expression and purification》2004,36(2):165-169
Tannase (tannin acyl hydrolase) is an industrially important enzyme produced by a large number of fungi, which hydrolyzes the ester and depside bonds of gallotannins and gallic acid esters. In the present work, a tannase from Aspergillus oryzae has been cloned and expressed in Pichia pastoris. The catalytic activity of the recombinant enzyme was assayed. A secretory form of enzyme was made with the aid of Saccharomyces cerevisiae alpha-factor, and a simple procedure purification protocol yielded tannase in pure form. The productivity of secreted tannase achieved 7000 IU/L by fed-batch culture. Recombinant tannase had a molecular mass of 90 kDa, which consisted of two kinds of subunits linked by a disulfide bond(s). Our study is the first report on the heterologous expression of tannase suggesting that the P. pastoris system represents an attractive means of generating large quantities of tannase for both research and industrial purpose. 相似文献
137.
Heather F. Armitage Andrea J. Britton René van der Wal Imogen S. K. Pearce Des B. A. Thompson Sarah J. Woodin 《Global Change Biology》2012,18(1):290-300
Ecosystems are subject to multiple, natural and anthropogenic environmental influences, including nitrogen (N) deposition, land use and climate. Assessment of the relative importance of these influences on biodiversity and ecosystem functioning is crucial for guiding policy and management decisions to mitigate global change; yet, few studies consider multiple drivers. In the UK, ongoing loss of the internationally important arctic/alpine moss‐sedge community, Racomitrium heath, has been linked to elevated N deposition, high grazing pressures and their combination; however, the relative importance of these drivers remains unclear. We used environmental gradients across the habitat's European distribution (UK, Faroes, Norway and Iceland) to investigate the relative impact of N deposition and grazing pressure, as well as climate, on the condition of the dominant moss species, Racomitrium lanuginosum. Key variables including tissue chemistry, growth and cover were measured at 36 sites, and multiple linear regressions were used to examine the relative importance of the drivers across sites. Our results clearly show that regional variation in the condition of R. lanuginosum across Europe is primarily associated with the impacts of N deposition, with climate (air temperature) and grazing pressure playing secondary roles. In contrast to previous experimental studies, we found moss growth to be stimulated by elevated N deposition; this apparent discrepancy may result from the use of artificially high N concentrations in many experiments. Despite increased growth rates, we found that moss mat depth and cover declined in response to N deposition. Our results suggest that this is due to increased decomposition of material in the moss mat, which ultimately leads to loss of moss cover and habitat degradation. This study clearly demonstrates both the key role of N deposition in degradation of Racomitrium heath and the importance of observational studies along natural gradients for testing predictions from experimental studies in the real world. 相似文献
138.
Qilei Wang Xiaojing Xue Yuling Li Yongbin Dong Long Zhang Qiang Zhou Fei Deng Zhiyan Ma Dahe Qiao Chunhui Hu Yangliu Ren 《Physiologia plantarum》2016,156(1):97-107
ADP‐ribosylation factors (ARFs) are small GTP‐binding proteins that regulate a wide variety of cell functions. Previously, we isolated a new ARF, ZmArf2, from maize (Zea mays). Sequence and expression characteristics indicated that ZmArf2 might play a critical role in the early stages of endosperm development. In this study, we investigated ZmArf2 function by analysis of its GTP‐binding activity and subcellular localization. We also over‐expressed ZmArf2 in Arabidopsis and measured organ and cell size and counted cell numbers. The expression levels of five organ size‐associated genes were also determined in 35S::ZmArf2 transgenic and wild‐type plants. Results showed that the recombinant ZmArf2 protein purified from Escherichia coli exhibited GTP‐binding activity. Subcellular localization revealed that ZmArf2 was localized in the cytoplasm and plasma membrane. ZmArf2 over‐expression in Arabidopsis showed that 35S::ZmArf2 transgenic plants were taller and had larger leaves and seeds compared to wild‐type plants, which resulted from cell expansions, not an increase in cell numbers. In addition, three cell expansion‐related genes, AtEXP3, AtEXP5 and AtEXP10, were upregulated in 35S::ZmArf2 transgenic lines, while the expression levels of AtGIF1 and AtGRF5, were unchanged. Collectively, our studies suggest that ZmArf2 has an active GTP‐binding function, and plays a crucial role in growth and development in Arabidopsis through cell expansion mediated by cell expansion genes. 相似文献
139.
Construction of a recombinant BHV-1 expressing the VP1 gene of foot and mouth disease virus and its immunogenicity in a rabbit model 总被引:1,自引:0,他引:1
Xian-Gang Ren Fei Xue Yuan-Mao Zhu Guang-Zhi Tong Yan-Hui Wang Jun-Ke Feng Hong-Fei Shi Yu-Ran Gao 《Biotechnology letters》2009,31(8):1159-1165
Foot-and-mouth disease (FMD) and infectious bovine rhinotracheitis (IBR) are two important infectious diseases of cattle.
Using bovine herpesvirus type 1 (BHV-1) as a gene delivery vector for development of live-viral vaccines has gained widespread
interest. In this study, a recombinant BHV-1 was constructed by inserting the synthetic FMDV (O/China/99) VP1 gene in the
the gE locus of BHV-1 genome under the control of immediately early gene promoter of human cytomegalovirus (phIE CMV) and
bovine growth hormone polyadenylation (BGH polyA) signal. After homologous recombination and plaque purification, a recombinant
virus named BHV-1/gE−/VP1 was acquired and identified. The immunogenicity was confirmed in a rabbit model by virus neutralization test and enzyme-linked
immunosorbent assay (ELISA). The result indicated that the BHV-1/gE−/VP1 has the potential for being developed as a bivalent vaccine for FMD and IBR. 相似文献
140.