嗜热脂肪芽孢杆菌DNA解链蛋白1的纯化及性质

以嗜热脂肪芽孢杆菌为材料,通过ＰｏｌｙｍｉｎＰ沉淀,硫酸铵分级及Ｐｈｅｎｙｌ－Ｓｅｐｈａｒｏｓｅ,ＤＥＡＥ纤维素,磷酸纤维素,ＦＰＬＣ ＭｏｎｏＱ,ＦＰＬＣ Ｓｕｐｅｒｏｓｅ１２等柱层析,得到部分纯化的ＤＮＡ解链蛋白１。ＢｓｔＨ１具有依赖ＤＮＡ和Ｍｇ＾２＋的ＡＴＰ酶活力,不同类型的核酸对ＢｓｔＨ１的ＡＴＰ酶活力的促进作用不同。     

限制性核酸内切酶Bsp 63Ⅰ的纯化及性质研究

用Ｂａｃｉｌｌｕｓｓｐｈａｅｒｉｃｕｓ６３菌为材料,经ＤＮＡ－Ｓｅｐｈａｒｏｓｅ和ＣｉｂａｃｒｏｎＢｌｕｅＦ３ＧＡ－Ｓｅｐｈａｒｏｓｅ两步亲和层析,将Ｂｓｐ６３Ⅰ纯化到均一程度。酶比活力达６１４００Ｕ／ｍｇ蛋白。用凝胶过滤法测得该酶分子量为１１３８００。该酶样品在ＳＤＳ－ＰＡＧＥ中呈现为一条蛋白带,并测得其亚基分子量为５６８００。用ＤＮＳ－Ｃｌ法测得该酶Ｎ－末端氨基酸为丙氨酸。上述结果表明该酶分子是由两个相同亚基组成。     

DOPE、DOPC对重组去脂肌质网Ca~(2+)-ATPase活力和构象的影响

经磷脂酶Ａ２ 去脂的肌质网Ｃａ２ ＋ － ＡＴＰａｓｅ 重组于不同比例的二油酰磷脂酰胆碱（Ｄｉｏｌｅｏｙｌｐｈｏｐｈａｔｉｄｙｌｃｈｏｌｉｎｅ,ＤＯＰＣ） 和二油酰磷脂酰乙醇胺（Ｄｉｏｌｅｏｙｌｐｈｏｐｈａｔｉｄｙｌｅｔｈａｎｏｌａｍｉｎｅ,ＤＯＰＥ） 形成脂酶体,研究了不同磷脂环境中Ｃａ２ ＋ － ＡＴＰａｓｅ 的ＡＴＰ 水解和Ｃａ２ ＋ 转运活力。结果表明,ＤＯＰＣ 和ＤＯＰＥ 分别有利于ＡＴＰ 水解和Ｃａ２ ＋ 的转运,ＤＯＰＥ 可以增强Ｃａ２ ＋ － ＡＴＰａｓｅ 的ＡＴＰ水解和Ｃａ２ ＋ 转运之间的偶联效率。利用内源荧光、荧光淬灭及Ｆｏｒｓｔｅｒ 能量转移原理测定Ｃａ２ ＋ －ＡＴＰａｓｅ 相应的构象变化, 发现随着ＤＯＰＥ／ ＤＯＰＣ 比例的改变使Ｃａ２ ＋ － ＡＴＰａｓｅ 构象发生相应的变化。     

甲基单胞菌Methylomonas sp.GYJ3中甲烷单加氧酶还原酶组分的纯化和性质

Ｍｅｔｙｌｏｍｏｎａｓｓｐ．ＧＹＪ３菌的甲烷单加氧酶（ＭＭＯ）粗酶提取液经ＤＥＡＥ－ＳｅｐｈａｒｏｓｅＣＬ－６Ｂ阴离子交换层析、ＳｅｐｈａｄｅｘＧ－１００凝胶过滤层析和ＤＥＡＥ－ＴＳＫｇｅｌＨＰＬＣ分离纯化出ＭＭＯ还原酶组分．经ＨＰＬＣ分析,纯度大于９５％,纯化倍数为４．４,加入至ＭＭＯ羟基化酶和调节蛋白Ｂ的体系中表现比活为２２８ｎｍｏｌ环氧丙烷每分钟毫克蛋白．ＳＤＳ－ＰＡＧＥ电泳表明还原酶由一种亚基组成,分子量４２ｋＤ．ＩＣＰ－ＡＥＳ测定还原酶的Ｆｅ含量为１．８３ｍｏｌＦｅ每ｍｏｌ蛋白．ＵＶ－Ｖｉｓ光谱表明还原酶除２８０ｎｍ蛋白质特征峰外在４６０ｎｍ有最大吸收峰,且Ａ２８０ｎｍ／Ａ４６０ｎｍ为２．５０,与其它黄素一铁硫蛋白相似,推测还原酶可能含一个ＦＡＤ辅基和Ｆｅ２Ｓ２中心．在厌氧条件下,还原酶能够和ＮＡＤＨ作用,ＵＶ－Ｖｉｓ光谱分析表明还原酶４６０ｎｍ处特征吸收峰消失,说明在ＭＭＯ催化过程中还原酶接受ＮＡＤＨ的电子．ＤＥＡＥ－ＳｅｐｈａｒｏｓｅＣＬ－６Ｂ阴离子交换层析分离出调节蛋白Ｂ,部分纯化的调节蛋白Ｂ的分子量大约在２０ｋＤ,它能够提高ＭＭＯ比活性４０倍,ＭＭＯ还原酶和调节蛋白Ｂ单独存在时不具有ＭＭＯ     

植物细胞质膜氧化还原系统

植物细胞质膜氧化还原系统陈珈（北京农业大学生物学院。北京１０００９４）ＲＥＤＯＸＳＹＳＴＥＭＩＮＴＨＥＰＬＡＳＭＡＭＥＭＢＲＡＮＥＯＦＰＬＡＮＴＣＥＬＬ￥ＣｈｅｎＪｉａ（ＣｏｌｌｅｇｅｏｆＢｉｏｌｏｇｉｃａ！Ｓｃｉｅｎｃｅｓ,ＢｅｉｊｉｎｇＡｇｒｉｃ...     

噬菌体RB69 DNA聚合酶的表达、分离纯化和其聚合反应的稳态动力学研究

噬菌体ＲＢ６９外切酶活性缺失的ＤＮＡ 聚合酶突变体（Ｄ２２２Ａ／Ｄ３２７Ａ）在大肠杆菌细胞中表达,表达量达细胞蛋白总量的６９％ ．表达后经ＤＥＡＥ－Ｓｅｐｈａｒｏｓｅ ＦａｓｔＦｌｏｗ , Ｓｏｕｒｃｅ ３０Ｑ 和ＨＴＰ三步分离纯化,纯度可达９９％ 以上．随后测定了该酶利用５种ｄＮＴＰ为底物进行聚合反应的酶促动力学常数（Ｋｍ 和Ｋｃａｔ）,结果表明该酶利用ｄＵＴＰ的能力与利用ｄＴＴＰ的能力相近,Ｋｍ （ｄＴＴＰ）和Ｋｍ（ｄＵＴＰ）均较高于其它３种脱氧核苷酸的Ｋｍ （ｄＡＴＰ, ｄＣＴＰ, ｄＧＴＰ）,推测其Ｋｍ 值的差异主要来源于Ｔ／Ｕ 碱基本身,而并非全部由ＧＣ碱基配对与ＡＴ碱基配对之间的氢键作用力的强弱差别所决定．     

高压快速柱分离纯化牛血球超氧化物歧化酶的研究

以牛血球为材料,经溶血等处理和丙酮沉淀,获得牛血球超氧化物歧化酶粗品。此粗酶可以通过ＤＥＡＥ－Ｓｅｐｈａｒｏｓｅ和ＣＭ－Ｓｅｐｈａｒｏｓｅ快速柱层析,获得超氧化物歧化酶纯品。纯化的酶比活可达１３５００ｕ／ｍｇ,经ＰＡＧＥ、ＳＤＳ－ＰＡＧＥ和快速蛋白液相色谱（ＦＰＬＣ）检测,结果表明,纯化酶是均一的Ｓｅｐｈａｄｅｘ Ｇ－１００凝胶过滤测得该酶分子量为３１,８００,ＳＤＳ－ＰＡＧＥ测得亚基分子量为１５     

大肠杆菌精氨酰—tRNA合成酶的变种ArgRS306KR的纯化…

本文从含ＡｒｇＲＳ３０６ＫＲ基因ａｒｇｓ３０６ＫＲ的ｐＵＣ１８重组质粒的大肠杆菌ＴＧ１转化子中经ＤＥＡＥ－Ｓｅｐｈａｃｅｌ和Ｂｌｕｅ－Ｓｅｐｈａｒｏｓｅ两步柱层析,得到电泳一条带的ＡｒｇＲＳ３０６ＫＲ。纯酶的比活为２７９０单位／毫克。该酶氨酰化和ＡＴＰ－ＰＰｉ交换活力的最适ＰＨ分别为ＰＨ８．３和ＰＨ７．５。氨酰化活力对ＡＴＰ、Ａｒｇ和ｔＲＮＡ的Ｋｍ分别２．６ｍｍｏｌ／Ｌ、１４．０μｍｏｌ／Ｌ和５．     

尖吻蝮蛇酸性磷脂酶A2的纯化和初步晶体学

为进一步揭示影响血小板聚集活性的结构基础,纯化了江西尖吻蝮蛇（Ａｇｋｉｓｔｒａｄｕｎ ａｃｕｔｕｓ）酸性磷脂酶Ａ２。江西省吻蝮蛇蛇毒经ＣＭ－Ｓｅｐｈａｒｏｓｅ离子交换柱层析,两步ＤＥＡＥ－Ｓｅｐｈａｒｏｓｅ离子交换柱层析以及Ｍｏｎｏ）ＧＦＰＬ离子交换柱层析,得到了ＳＤＳ聚丙烯酰胺凝胶电泳和等和集电泳均一的磷脂酶Ａ２（ＰＬＡ２）,其分子量为１６．５ＫＤ,等电点为４．３,经测定,该酶具有抑制ＡＤＰ诱地     

抗癌胚抗原单链抗体基因在家蚕细胞和幼虫中的表达

将抗癌胚抗原（ＣＥＡ）单链抗体基因插入家蚕杆状病毒转移载体ｐＢａｃＰＡＫＨｉｓ, 与修饰的家蚕核型多角体病毒ＢｍＢａｃＰＡＫＤＮＡ共转染家蚕细胞, 经同源重组得到含有在多角体蛋白基因启动子控制下的抗ＣＥＡＳｃＦｖ 基因的重组病毒ＢｍＢａｃＳｃＦｖ。用重组病毒分别感染家蚕细胞和幼虫, 在两者中均得到了高效表达, 产物分子量为２８ｋＤ, 前者占细胞总蛋白的６ ％ , 后者为０ ．３ ｍｇ／ 蚕。目的基因在家蚕细胞和幼虫中表达产物经Ｎｉ２＋ＩＤＡＳｅｐｈａｒｏｓｅ６Ｂ亲和柱纯化, 前者纯度可达９０％ 以上, 后者纯度较低; 纯化后的融合蛋白具有ＣＥＡ 结合活力, 其亲和常数分别为５ ．４×１０８／ｍｏｌ·Ｌ－ １ 和２．３ ×１０８／ｍｏｌ·Ｌ－１ , 略低于其亲本单抗Ｅ７Ｂ１０ ２．７ ×１０９／ｍｏｌ·Ｌ－ １ 。     

A Unique Protease Cleavage Site Predicted in the Spike Protein of the Novel Pneumonia Coronavirus (2019-nCoV) Potentially Related to Viral Transmissibility

正Dear Editor,In December 2019, a novel human coronavirus caused an epidemic of severe pneumonia(Coronavirus Disease 2019,COVID-19) in Wuhan, Hubei, China(Wu et al. 2020; Zhu et al. 2020). So far, this virus has spread to all areas of China and even to other countries. The epidemic has caused 67,102 confirmed infections with 1526 fatal cases     

Curcuminoids as inhibitors of thioredoxin reductase: A receptor based pharmacophore study with distance mapping of the active site

Curcumin is the yellow pigment of turmeric that interacts irreversibly forming an adduct with thioredoxin reductase (TrxR), an enzyme responsible for redox control of cell and defence against oxidative stress. Docking at both the active sites of TrxR was performed to compare the potency of three naturally occurring curcuminoids, namely curcumin, demethoxy curcumin and bis-demethoxy curcumin. Results show that active sites of TrxR occur at the junction of E and F chains. Volume and area of both cavities is predicted. It has been concluded by distance mapping of the most active conformations that Se atom of catalytic residue SeCYS498, is at a distance of 3.56 from C13 of demethoxy curcumin at the E chain active site, whereas C13 carbon atom forms adduct with Se atom of SeCys 498. We report that at least one methoxy group in curcuminoids is necessary for interation with catalytic residues of thioredoxin. Pharmacophore of both active sites of the TrxR receptor for curcumin and demethoxy curcumin molecules has been drawn and proposed for design and synthesis of most probable potent antiproliferative synthetic drugs.     

Comparative morphology of the carpel in the Liliaceae: Hewardieae, Petrosavieae, and Tricyrteae

The young pistils in the melanthioid tribes, Hewardieae, Petrosavieae and Tricyrteae, are uniformly tricarpellate and syncarpous. They lack raphide idioblasts. All are multiovulate, with bitegmic ovules. The Petrosavieae are marked by the presence of septal glands and incomplete syncarpy. Tepals and stamens adhere to the ovary in the Hewardieae and the Petrosavieae but not in the Tricyrteae. Two vascular bundles occur in the stamens of the Hewartlieae and Tricyrtis latifolia. Ventral bundles in the upper part of the ovary of the Hewardieae are continuous with compound septal bundles and placental bundles in the lower part. Putative ventral bundles occur in the alternate position in the Tricyrteae and putative placental bundles in the opposite. position in the Petrosavieae. The dichtomously branched stigma in each carpel of the Tricyrteae is supplied by a bifurcated dorsal bundle.     

Enterovirus 71 3C proteolytically processes the histone H3 N-terminal tail during infection

Highlights
1. The N-terminal tail of histone H3 is specifically cleaved during EV71 infection.
2. Viral protease 3C is identified as a protease responsible for proteolytically processing the N-terminal H3 tail.
3. Our finding reveals a new epigenetic regulatory mechanism for Enterovirus 71 in virus-host interactions.     

Detection of EBV and HHV6 in the Brain Tissue of Patients with Rasmussen’s Encephalitis

Rasmussen’s encephalitis (RE) is a rare pediatric neurological disorder, and the exact etiology is not clear. Viral infection may be involved in the pathogenesis of RE, but conflicting results have reported. In this study, we evaluated the expression of both Epstein-Barr virus (EBV) and human herpes virus (HHV) 6 antigens in brain sections from 30 patients with RE and 16 control individuals by immunohistochemistry. In the RE group, EBV and HHV6 antigens were detected in 56.7% (17/30) and 50% (15/30) of individuals, respectively. In contrast, no detectable EBV and HHV6 antigen expression was found in brain tissues of the control group. The co-expression of EBV and HHV6 was detected in 20.0% (6/30) of individuals. In particular, a 4-year-old boy had a typical clinical course, including a medical history of viral encephalitis, intractable epilepsy, and hemispheric atrophy. The co-expression of EBV and HHV6 was detected in neurons and astrocytes in the brain tissue, accompanied by a high frequency of CD8⁺ T cells. Our results suggest that EBV and HHV6 infection and the activation of CD8⁺ T cells are involved in the pathogenesis of RE.     

Perinatal Vertical Transmission of Chikungunya Virus in Ruili,a Town on the Border between China and Myanmar

正Dear Editor,Chikungunya virus (CHIKV), an arbovirus in the family of Togaviridae, genus Alphavirus, is transmitted by the A.aegyptii or A. albopictus mosquito, and causes disease in humans characterized by fever, rash, and arthralgia (Silva and Dermody 2017; Suhrbier 2019). It was first reported in 1953 in Tanzania, and caused only a few outbreaks and sporadic cases in Africa and Asia in last century. However, in the epidemic in 2004, CHIKV acquired mutations that conferred enhanced transmission by the A. albopictus mosquito(Schuffenecker et al. 2006). Since then, it has successively caused outbreaks in Africa, the Indian Ocean, South East Asia, the South America, and Europe (Zeller et al. 2016).     

Development of a Novel Reverse Transcription Loop-Mediated Isothermal Amplification Method for Rapid Detection of SARS-CoV-2

In conclusion, the novel visual RT-LAMP assay is a simple, rapid, and sensitive approach for detection of SARS-CoV-2, and it is ready for application in primary care and community hospitals or health care centers, and even patients' own houses in response to the current SARS-CoV-2 epidemic because the assay does not require sophisticated equipment and skilled personnel. Furthermore, it is also ready to be used in fields for screening samples from wild animals and environments to facilitate the identification of potential intermediate hosts that mediate the cross-species transmission of SARS-CoV-2 from bats to humans.