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附属蛋白:冠状病毒不容忽视的一类蛋白 被引量:2
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作者 司伏生 姜黎 +4 位作者 于瑞嵩 董世娟 谢春芳 陈冰清 李震 《微生物学通报》 CAS CSCD 北大核心 2021年第10期3895-3909,共15页
冠状病毒(Coronaviruses,CoVs)是基因组最大的一类单股正链RNA病毒,多数可以跨物种传播并感染人类,是当前引起重大公共卫生事件、严重威胁人类健康的病原之一。病毒基因组全长约25-31 kb,编码多个非结构蛋白、结构蛋白(S、E、M、N)及附... 冠状病毒(Coronaviruses,CoVs)是基因组最大的一类单股正链RNA病毒,多数可以跨物种传播并感染人类,是当前引起重大公共卫生事件、严重威胁人类健康的病原之一。病毒基因组全长约25-31 kb,编码多个非结构蛋白、结构蛋白(S、E、M、N)及附属蛋白。对于大多数冠状病毒来说,附属蛋白虽然是病毒复制的非必需蛋白,但往往在病毒的致病过程中发挥重要作用,是冠状病毒重要的功能蛋白。该类蛋白位于病毒基因组的3′端,由位于基因起始位置的转录调控序列(Transcription Regulating Sequence,TRS)调控其m RNA的转录,而且蛋白编码序列的密码子使用偏爱性对蛋白翻译也产生重要影响。附属蛋白具有跨膜蛋白的属性和独特的蛋白转运基序,后者对该类蛋白跨膜区的形成、拓扑学结构及蛋白的细胞内运输过程起决定性的作用,从而直接影响附属蛋白的功能。本文首先总结了冠状病毒最新的分类及基因组结构;然后从附属蛋白的种类、功能、蛋白转运基序、拓扑学结构及密码子使用偏爱性等方面系统概述了相关研究进展,并对下一步的研究方向进行了展望,为更加全面地认识冠状病毒附属蛋白的生物学特性提供重要参考。 展开更多
关键词 冠状病毒 附属蛋白 细胞内转运 拓扑学 密码子使用偏爱性
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猪流行性腹泻病毒经其附属蛋白抑制细胞凋亡 被引量:2
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作者 王晨阳 司伏生 +4 位作者 于瑞嵩 谢春芳 董世娟 宋增福 李震 《微生物学通报》 CAS CSCD 北大核心 2020年第8期2484-2494,共11页
【背景】orf3位于猪流行性腹泻病毒(porcine epidemic diarrhea virus,PEDV)s基因与e基因之间,是目前发现的PEDV唯一一个附属基因,编码附属蛋白(ORF3蛋白)。我们前期研究初步发现ORF3蛋白对PEDV诱导的细胞凋亡有影响。【目的】研究ORF3... 【背景】orf3位于猪流行性腹泻病毒(porcine epidemic diarrhea virus,PEDV)s基因与e基因之间,是目前发现的PEDV唯一一个附属基因,编码附属蛋白(ORF3蛋白)。我们前期研究初步发现ORF3蛋白对PEDV诱导的细胞凋亡有影响。【目的】研究ORF3蛋白在PEDV侵染复制过程中的毒力作用机制。【方法】实验用3种PEDV:rDR13att-∆ORF3(orf3基因全部敲除)、DR13-ORF3att(携带有C端截短orf3)、rDR13att-ORF3wt(携带全长orf3基因)感染Vero细胞,观察病变情况,再用活细胞成像仪、流式细胞仪、DNA断裂的原位末端标记法[terminal deoxynucleotidyl transferase(TDT)-mediated dUTP nick end labeling,TUNEL]等方法检测不同感染时间点的细胞凋亡情况,然后用蛋白质印迹方法分析PEDV感染宿主细胞中主要凋亡相关蛋白(如Caspase-3)的活化或裂解,最后进行转录组测序研究病毒感染细胞中差异基因的表达情况,再用荧光定量PCR验证转录组结果。【结果】rDR13att-∆ORF3引起较多的细胞病变,活细胞成像仪的动态观察结果显示,3种病毒侵染的细胞凋亡水平随着时间的延长均高于正常阴性细胞,但敲除orf3的病毒感染细胞后细胞凋亡率比其他两种病毒更高;敲除orf3病毒感染细胞凋亡率显著高于其他两种病毒;病毒rDR13att-∆ORF3感染细胞后TUNEL阳性细胞数比DR13-ORF3att和rDR13att-ORF3wt更多;表达ORF3蛋白的重组PEDV可以抑制Caspase-3的活化;ORF3蛋白对受感染细胞Heat shock 70 kD protein 1B(HSP70)基因转录有促进作用,荧光定量PCR结果表明rDR13att-ORF3wt感染细胞的HSP70表达量高于rDR13att-∆ORF3感染细胞。【结论】PEDV通过ORF3蛋白抑制细胞凋亡,而且这种作用可能是通过抑制Caspase-3的活化或增加HSP70的产生来完成的。 展开更多
关键词 猪流行性腹泻病毒 附属ORF3蛋白 功能 抑制 细胞凋亡
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心跳紊乱之谜
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《发明与创新(大科技)》 2003年第8期43-43,共1页
关键词 心电紊乱 QT延长综合症 附属蛋白蛋白B 基因缺陷 心脏疾病 病因
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Adsorption of Heavy Metal Ions,Dyes and Proteins by Chitosan Composites and Derivatives-A Review 被引量:16
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作者 LIU Bingjie WANG Dongfeng +1 位作者 YU Guangli MENG Xianghong 《Journal of Ocean University of China》 SCIE CAS 2013年第3期500-508,共9页
Chitosan composites and derivatives have gained wide attentions as effective biosorbents due to their low costs and high contents of amino and hydroxyl functional groups.They have showed significant potentials of remo... Chitosan composites and derivatives have gained wide attentions as effective biosorbents due to their low costs and high contents of amino and hydroxyl functional groups.They have showed significant potentials of removing metal ions,dyes and proteins from various media.Chemical modifications that lead to the formation of the chitosan derivatives and chitosan composites have been extensively studied and widely reported in literatures.The aims of this review were to summarize the important information of the bioactivities of chitosan,highlight the various preparation methods of chitosan-based active biosorbents,and outline its potential applications in the adsorption of heavy metal ions,dyes and proteins from wastewater and aqueous solutions. 展开更多
关键词 chitosan composite chitosan derivative ADSORPTION heavy metal DYE protein
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A NOVEL METAL CHELATE AFFINITY ADSORBENT FOR PROTEIN UPTAKE
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作者 WANG Yongjian BAI Shu SUN Yan 《Chinese Journal of Reactive Polymers》 2001年第2期134-140,共7页
In this article, a spherical chitosan gel crosslinked by epichlorohydrin was prepared. It was then loaded with copper ions to produce a metal chelate affinity adsorbent for protein. The uptake of bovine serum albumin ... In this article, a spherical chitosan gel crosslinked by epichlorohydrin was prepared. It was then loaded with copper ions to produce a metal chelate affinity adsorbent for protein. The uptake of bovine serum albumin (BSA) by the affinity adsorbent was investigated, and the adsorption capacity for BSA as high as 40 mg/g-wet beads was observed. The adsorption equilibrium data was well correlated by the Langmuir equation. The adsorption was considerably affected by pH. In addition, the amount of BSA adsorbed onto the beads decreased with the increasing of aqueous phase ionic strength, so adsorbed BSA can be desorbed by adjusting pH or ionic strength of the solution. 展开更多
关键词 CHITOSAN EPICHLOROHYDRIN BSA Bounded copper Adsorption
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Autoantibodies against MMP-7 as a novel diagnostic biomarker in esophageal squamous cell carcinoma 被引量:8
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作者 Jing-Hua Zhou Bin Zhang +1 位作者 Kemp H Kernstine Li Zhong 《World Journal of Gastroenterology》 SCIE CAS CSCD 2011年第10期1373-1378,共6页
AIM: To evaluate the diagnostic values of serum autoan tibodies against matrix metalloproteinase-7 (MMP-7) in patients with esophageal squamous cell carcinoma (ESCC). METHODS: The MMP-7 cDNA was cloned from ESCC... AIM: To evaluate the diagnostic values of serum autoan tibodies against matrix metalloproteinase-7 (MMP-7) in patients with esophageal squamous cell carcinoma (ESCC). METHODS: The MMP-7 cDNA was cloned from ESCC tissues, and MMP-7 was expressed and purified from a prokaryotic system. MMP-7 autoanUbodies were then measured in sera from 50 patients with primary ESCC and 58 risk-matched controls, using a reverse capture enzyme-linked immunosorbent assay (ELISA) in which autoantibodies to MMP-7 bound to the purified MMP-7 proteins. In addition, MMP-7 autoantibody levels in sera from 38 gastric cancer patients and from control serum samples were also tested. RESULTS: The optimum conditions for recombinant MMP-7 protein expression were determined as 0.04 mmol/L Isopropyl-13-D-Thiogalactopyranoside (IPTG)induction at 37℃ for four hours. The levels of serum autoantibodies against MMP-7 were significantly higher in patients with ESCC than in the matched-control samples (OD450 = 1.69 ±0.08 vs OD450 = 1.55 ± 0.10, P 〈 0.001). The area under the receiver operating character- istic (ROC) curve was 0.87. The sensitivity and specificity for detection of ESCC were 78.0% and 81.0%, respectively, when the OD450 value was greater than 1.65. Although the levels of autoantibodies against MMP-7 were also significantly higher in patients with gastric cancer compared to control samples (OD450 = 1.62± 0.06 vs OD450 = 1.55±0.10, P 〈 0.001), the diagnostic accuracy was less significant than in ESCC patients. The area of ROC curve was 0.75, whereas the sensitivity and specificity were 60.5% and 71.7%, respectively, when the cut-off value of OD450 was set at 1.60. CONCLUSION: Serum autoantibody levels of MMP-7 may be a good diagnostic biomarker for esophageal squamous cell carcinoma, 展开更多
关键词 Matrix metalloproteinase-7 Serum autoantibody Esophageal squamous cell carcinoma Gastriccancer BIOMARKER
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中东呼吸综合征冠状病毒ORF5及其突变型基因真核表达载体的构建与表达
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作者 张世豪 黄姗 +5 位作者 陈嘉仪 高静 蔡金泰 杨秀文 赵卫 张宝 《中国病原生物学杂志》 CSCD 北大核心 2021年第5期507-512,519,共7页
目的对中东呼吸综合征冠状病毒(MERS-CoV)ORF5及其突变型基因进行真核表达载体的构建与表达,为进一步研究ORF5作用机制奠定基础。方法将MERS-CoV野生ORF5基因序列(KT036372),截短ORF5(ORF5△)基因序列(KT036373)以及融合ORF5(ORF5+)基... 目的对中东呼吸综合征冠状病毒(MERS-CoV)ORF5及其突变型基因进行真核表达载体的构建与表达,为进一步研究ORF5作用机制奠定基础。方法将MERS-CoV野生ORF5基因序列(KT036372),截短ORF5(ORF5△)基因序列(KT036373)以及融合ORF5(ORF5+)基因序列(KT036374)克隆至真核表达载体质粒pEGFP-N1中,构建目的基因与EGFP融合表达质粒。采用定点突变技术在ORF5与EGFP之间添加终止密码或移码突变,并将ORF5、ORF5△、ORF5+蛋白126位、136位起始密码子突变为终止密码子。将测序正确的质粒转染HEK-293T细胞,Western blot检测相应蛋白的表达情况。结果成功构建ORF5、ORF5△、ORF5+真核表达载体。ORF5与EGFP融合蛋白在荧光显微镜下可见强荧光,但Western blot仅能检出EGFP蛋白,未能检出完整的融合蛋白;当ORF5与EGFP之间加入终止密码或移码突变后,仍能够检出EGFP蛋白。ORF5△、ORF5+与EGFP融合蛋白成功表达,以EGFP为抗原,ORF5△、ORF5+可检出多种不同分子质量蛋白,126位终止后仍可检出多种不同分子质量蛋白,136位终止后ORF5△缺少1种目的蛋白。结论成功构建MERS-CoV野生、截短以及融合ORF5的真核表达质粒。野生ORF5蛋白表达机制复杂,该蛋白可能存在类似内部核糖体进入位点(internal ribosome entry site,IRES)的特殊功能。变异体ORF5的表达存在多个翻译起始位点,其作用机制及功能需进一步研究。 展开更多
关键词 中东呼吸综合征冠状病毒 附属蛋白 真核表达载体构建 Western blot
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Transforming growth factor-α promotes mouse blastocyst outgrowth and secretion of matrix metalloproteinases
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作者 陈舒怡 曹宇静 +1 位作者 曾国庆 段恩奎 《Chinese Medical Journal》 SCIE CAS CSCD 2001年第12期68-72,108,共6页
Objective To study the effect of transforming growth factor-α (TGF-α) on early stage of embryo implantation.Methods Mouse blastocysts were cultured in vitro in medium containing various concentrations of TGF-α. Bla... Objective To study the effect of transforming growth factor-α (TGF-α) on early stage of embryo implantation.Methods Mouse blastocysts were cultured in vitro in medium containing various concentrations of TGF-α. Blastocyst implantation capacity was evaluated by calculating the percentage of embryos with attachment or outgrowth. Matrix metalloproteinases (MMPs) secretion of blastocysts was observed using gelatin zymography. Results There was no significant difference in the percentage of attachment between control and TGF-α treated groups, but the percentage of outgrowth of TGF-α treated groups was significantly higher than that of the control group after 24h culturing. Gelatin zymography showed that blastocysts cultured in TGF-α treated groups started secreting MMPs earlier than those in the control group.Conclusion TGF-α is involved in regulating the mouse embryo implantation process by promoting blastocyst outgrowth and secreting matrix matalloproteinases. 展开更多
关键词 transforming growth factor-α · embryo implantation · attachment · outgrowth · matrix metalloproteinases
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