辛德毕斯病毒装配及其6K蛋白与中间纤维的关系

用温和的选择性抽提方法与整装细胞电镜技术,DGD包埋-去包埋剂电镜技术相结合,对辛德毕斯病毒的装配与宿主细胞中间纤维的关系进行了探讨。电镜观察清晰地显示了病毒“装配中心”被中间纤维所网络,病毒的装配过程显然是以中间纤维为支架;正在装配的核壳体与已装配的核壳体紧密结合在中间纤维丝上。根据电镜照片分析,核壳体可能是沿中间纤维由装配中心向外扩散。应用人工合成6K蛋白所得抗体进行胶体金标记,证明辛德毕斯病毒非结构性6K蛋白也与中间纤维紧密结合。     

化脓性链球菌纤维粘连蛋白结合蛋白的研究进展

本文介绍化脓性链球菌纤维粘连蛋白（ＥＢＰ）结合蛋白的最新研究进展，对链球菌的粘附机理、纤维粘连蛋白结合蛋白的分子生物学结构特征、在粘附过程中的作用以及其临床意义作了扼要阐述。     

人疱疹病毒6型基因编码蛋白研究进展

人疱疹病毒6 型( HHV-6) 是一类嗜人淋巴细胞的双链DNA 病毒, 属疱疹病毒β亚科。研究推测HHV-6 基因组可编码80 ～100 种蛋白质, 包括膜蛋白、即刻早期( IE) 蛋白、DNA 蛋白、DNA 包装蛋白、病毒装配蛋白等。本文就HHV-6 基因编码蛋白的特性和功能研究进展予以综述。     

钙连蛋白——膜结合型监护蛋白

钙连蛋白──膜结合型监护蛋白许强,王克夷（中国科学院上海生物化学研究所,上海２０００３１）关键词钙连蛋白,监护蛋白１．发现内质网（ＥＲ）是许多分泌蛋白、质膜蛋白和细胞器膜蛋白加工处理的场所,这些蛋白在ＥＲ经过折叠、装配和修饰（例如Ｎ－糖基化）以后,才...     

肌动蛋白集束调控因子及G蛋白信号转导通路

细胞骨架由微丝、微管及中等纤维组成受不同蛋白因子调控以不同方式组装成不同直径的纤维 ,遍布于一切细胞 ,决定细胞的形状 ,赋予其抗压强度 ,对细胞器及大分子进行空间组织 ,实现胞内的能量转换。在肌动蛋白 (ａｃｔｉｎ)组装成张力纤维和张力纤维解离成肌动蛋白单体过程中有多种蛋白因子参与调控 ,从而使细胞骨架处于一个生理的动态平衡中 ,执行和完成不同的生化反应。在众多的调控蛋白中 ,肌动蛋白集束调控蛋白因子 (ａｃｔｉｎｂｕｎｄｌｉｎｇｐｒｏｔｅｉｎ)不仅参与肌动蛋白结构调节 ,还与细胞内信号传导有密切关系。已发现的肌动蛋…     

蜘蛛丝蛋白研究进展

由于蜘蛛丝蛋白分子高度重复的一级结构、特殊的溶解特性和分子折叠行为以及具有形成非凡力学特性丝纤维的能力而引人注目。本文从蛛丝蛋白基因、天然蛛丝形成过程、蛛丝蛋白的基因工程生产及蛛丝蛋白的应用前景等几个方面着重介绍了近20年来对蛛丝蛋白的研究进展。围绕蛛丝蛋白展开的研究将有助于揭示蛋白质一级结构、蛋白质分子折叠与蛋白质大分子特性之间的内在联系。     

热休克蛋白的生物学功能

热休克蛋白是所有原核细胞和真核细胞遭受高温或其他应激所产生一组非常保守的蛋白分子家族。它们介导其他蛋白质的跨膜运和正确装配,起着分子伴侣的重要的作用;参与甾体激素受体等的功能发挥,与细胞的许多重要生理过程相关;在肿瘤细胞中,参与癌基因蛋白ＰＰ６０＾ｓｒｃ及抗癌基因蛋白Ｐ５３的作用,并结合核内蛋白改变其功能作用。     

检测尿液中纤维结合蛋白可以帮助发现膀胱癌

据西班牙研究人员说,测量尿液中的纤维结合蛋白可能帮助诊断膀胱癌。研究者发现,纤维结合蛋白使用25．6mg／L的界值、纤维结合蛋白与肌酐之比使用36．9的界值对膀胱癌分别有78％和75％的敏感性,特异性都是80％。尿细胞学的敏感性则只有55％,但特异性达100％。研究者还发现纤维结合蛋白、纤维结合蛋白与肌酐之比根据肿瘤的大小和阶段而显著不同,他们认为这些结果可能对制订治疗决策有意义。     

两个棉花Rac蛋白基因的克隆与表达分析

为研究棉花纤维起始和伸长的分子机理,在棉花纤维EST序列分析的基础上,从棉花纤维中扩增并克隆了2个棉花Rac蛋白的cDNA基因,分别命名为GhRacA和GhRacB。GhRacA cDNA长959bp,推测的编码蛋白包含211个氨基酸。GhRacB cDNA长920bp,编码195个氨基酸的蛋白。GhRacA和GhRacB蛋白均含有GTP／GDP结合和激活区域、Effector区和碱性氨基酸区。GhRacB的C末端有保守的异戊烯基化位点CSIL,而GhRacA没有明显的异戊烯基化位点。序列比较分析表明,GhRacA和GhRacB是2个新的棉花Rac蛋白。RT-PCR分析表明,GhRacA和GhRacB在根、下胚轴、茎、叶和纤维中都有表达,但均在棉花纤维起始和伸长时期有优势表达,推测2个基因在棉花纤维的早期发育中可能有重要的功能。     

DnaJ—like蛋白研究进展

ＤｎａＪ－ｌｉｋｅ蛋白由Ｎ－端保守的Ｊ区域、富含Ｇｌｙ和Ｐｈｅ区域,富含Ｇｙｓ区域和Ｃ－端低同源区域组成。Ｊ功能域能调节ＨＳＰ７０分子伴侣的ＡＴＰａｓｅ活性,Ｃ－端不保守域能调节与多肽的关系。直核细胞中存在着多种结构不同的ＤｎａＪ－ｌｉｋｅ蛋白,但都含有一个Ｊ功能域。ＤｎａＪ－ｌｉｋｅ蛋白通过Ｊ功能域调节ＨＳＰ７０功能而参与蛋白的折叠,装配和运输过程。     

Innervation of periodontal ligament and dental pulp in the rat incisor: An immunohistochemical investigation of neurofilament protein and glia-specific S-100 protein

Summary Nervous elements in the periodontal ligament and dental pulp of rat incisors were investigated by means of immunohistochemistry for neurofilament protein (NFP) and glia-specific S-100 protein. The periodontal ligament in the incisors was densely innervated by NFP-immunoreactive nerve fibers; the distribution of the nerve fibers and their terminations differed markedly from those in molars. NFP-positive, thick nerve bundles entered the lingual periodontal ligament through slits located in the mid-region of the alveolar socket, and immediately formed numerous Ruffini-like corpuscles. In the labial periodontal ligament, all of the NFP-immunoreactive nerve fibers terminated in free endings. The restricted location of the stretch receptor, Ruffini-like corpuscle, in the lingual periodontal ligament appears to be an essential element, because this region is regularly extended during mastication. The nervous elements were restricted to the alveolar half of the periodontal ligament in every region; they avoided the dental half of the periodontal ligament, which presumably moves continuously with the tooth. Pulpal nerve fibers in incisors also showed a characteristic distribution different from those in molars; individual nerve fibers with beaded structures ran in the center of the pulp toward the incisai edge, and did not form the subodontoblastic nerve plexus of Raschkow.Immunostaining for S-100 protein revealed a distribution pattern of nervous elements similar to that for NFP, suggesting that the nerves supplying the periodontal ligament and dental pulp were mostly covered by a Schwann sheath.     

beta-Amyloid peptide induces formation of actin stress fibers through p38 mitogen-activated protein kinase

Based on the critical role of actin in the maintenance of synaptic function, we examined whether expression of familial beta-amyloid precursor protein APP-V642I (IAPP) or mutant presenilin-1 L286V (mPS1) affects actin polymerization in rat septal neuronal cells. Expression of either IAPP or mPS1 but not wild-type amyloid precursor protein or presenilin-1induced formation of actin stress fibers in SN1 cells, a septal neuronal cell line. Treatment with beta-amyloid (Abeta) peptide also caused formation of actin stress fibers in SN1 cells and primary cultured hippocampal neurons. Treatment with a gamma-secretase inhibitor completely blocked formation of actin stress fibers, indicating that overproduction of Abeta peptide induces actin stress fibers. Because activation of the p38 mitogen-activated protein kinase (p38MAPK)-mitogen-associated protein kinase-associated protein kinase (MAPKAPK)-2-heat-shock protein 27 signaling pathway mediates actin polymerization, we explored whether Abeta peptide activates p38MAPK and MAPKAPK-2. Expression of IAPP or mPS1 induced activation of p38MAPK and MAPKAPK-2. Treatment with a p38MAPK inhibitor completely inhibited formation of actin stress fibers mediated by Abeta peptide, IAPP or mPS1. Moreover, treatment with a gamma-secretase inhibitor completely blocked activation of p38MAPK and MAPKAPK-2. In summary, our data suggest that overproduction of Abeta peptide induces formation of actin stress fibers through activation of the p38MAPK signaling pathway in septal neuronal cells.     

棉花纤维蛋白质3种提取及二维电泳方法的比较

高质量的蛋白样品制备是进行二维电泳的先决条件.棉花纤维中含有纤维素、多酚、多糖等严重干扰二维电泳的物质, 增加了蛋白提取和二维电泳的难度.分别采用3种提取植物组织蛋白的方法(水法、酚法和尿素法), 提取棉纤维总蛋白, 进而进行了二维电泳分析.在蛋白产量、蛋白纯度和电泳图谱等方面对3种方法进行了比较, 结果采用酚法提取的样品取得了较好的电泳图谱, 有望成为从棉纤维样品中提取总蛋白的可选方法.     

突触体素、S-100蛋白、NSE免疫反应神经纤维在人淋巴结的分布

探讨突触体素、S－100蛋白、NSE免疫反应神经纤维在人淋巴结的分布,为淋巴结的神经免疫相互作用提供形态学资料。应用免疫组织化学ABC法观察人类腹股沟、腋窝、肠系膜、肺等淋巴结40例,10％福尔马林固定,石蜡包埋组织切片。结果显示：突触体素、S－100蛋白、NSE免疫反应神经纤维呈细丝状沿被膜和门部结缔组织小梁及血管进入皮质后主要分布于副皮质区,环境淋巴小结,进一步分支到达髓质。同时在淋巴小结发生中心及副皮质区有S－100蛋白免疫反应阳性细胞。在髓质髓窦内有NSE免疫反应阳性细胞。结论;淋巴结内有突触体素、S－100蛋白、NSE免疫反应神经纤维的支配、并有S－100蛋白、NSE免疫反应阳性细胞,为淋巴结的神经免疫相互作用提供形态学资料。     

Microinjection of Protein Tyrosine Phosphatases into Fibroblasts Disrupts Focal Adhesions and Stress Fibers

Microinjection and scrape-loading have been used to load cells in culture with soluble protein tyrosine phosphatases (FTPs). The introduction of protein tyrosine phosphatases into cells caused a rapid (within 5 minutes) decrease in tyrosine phosphorylation of major tyrosine phosphorylated substrates, including the focal adhesion kinase and paxillin. This decrease was detected both by blotting whole cell lysates with anti-phosphotyrosine antibodies and visualizing the phosphotyrosine in focal adhesions by immunofluorescence microscopy. After 30 minutes, many of the cells injected with tyrosine phosphatases revealed disruption of focal adhesions and stress fibers. To determine whether this disruption was due to the dephosphorylation of FAK and its substrates in focal adhesions, we have compared the effects of protein tyrosine phosphatase microinjection with the effects of displacing FAK from focal adhesions by microinjection of a dominant negative FAK construct. Although both procedures resulted in a marked decrease in the level of phosphotyrosine in focal adhesions, disruption of focal adhesions and stress fibers only occurred in cells loaded with exogenous protein tyrosine phosphatases. These results lead us to conclude that although tyrosine phosphorylation regulates focal adhesion and stress fiber stability, this does not involve FAK nor does it appear to involve tyrosine-phosphorylated proteins within focal adhesions. The critical tyrosine phosphorylation event is upstream of focal adhesions, a likely target being in the Rho pathway that regulates the formation of stress fibers and focal adhesions.     

Identification and localization of a protein immunologically related to Caltractin (Centrin) in the Myonemes and Membranelles of the Heterotrich ciliate Stentor coeruleus

The contractile properties of the myonemes of Stentor are very similar to caltractin (centrin)-containing fibers of other organisms. We investigated whether the calcium-binding protein caltractin was present in Stentor by using three different antibodies to caltractin or caltractin-related proteins, in conjunction with immunofluorescence microscopy and protein blotting. Immunofluorescence demonstrated that a protein immunologically similar to caltractin is present in the myonemes and in the bases of the membranelles of Stentor. The localization to the myonemes is observed in intact cells, osmotically lysed cells, and isolated cortices. Double-label immunofluorescence with anti-alpha-tubulin and anti-caltractin antibodies showed that the fluorescence in the myonemes was not in the overlying Km fibers. The myonemes in the posterior one-third of the cell appear as thick fibers with no cross-bridging. They become thinner as they approach the anterior end of the cell and show extensive cross-bridging here. Staining in the bases of the membranelles shows a distinct comma-like immunofluorescence pattern similar to that seen with protargol-stained cells and SEM views of the membranellar band reported by others. Western blots demonstrated that the caltractin-like protein in Stentor has an apparent molecular weight of 23 kDa compared with the 20-kDa protein from Chlamydomonas and is a calcium-binding protein.     

Homocysteine-induced endoplasmic reticulum protein (Herp) is up-regulated in sporadic inclusion-body myositis and in endoplasmic reticulum stress-induced cultured human muscle fibers

Herp is a stress-response protein localized in the endoplasmic reticulum (ER) membrane. Herp was proposed to improve ER-folding, decrease ER protein load, and participate in ER-associated degradation (ERAD). Intra-muscle-fiber ubiquitinated multiprotein-aggregates containing, among other proteins, either amyloid-beta (Abeta) or phosphorylated tau are characteristic of sporadic inclusion-body myositis (s-IBM). ER stress and proteasome inhibition appear to play a role in s-IBM pathogenesis. We have now studied Herp in s-IBM muscle fibers and in ER-stress-induced or proteasome-inhibited cultured human muscle fibers. In s-IBM muscle fibers: (i) Herp was strongly immunoreactive in the form of aggregates, which co-localized with Abeta, GRP78, and beta2 proteasome subunit; (ii) Herp mRNA and protein were increased. In ER-stress-induced cultured human muscle fibers: (i) Herp immunoreactivity was diffusely increased; (ii) Herp mRNA and protein were increased. In proteasome-inhibited cultured human muscle fibers: (i) Herp immunoreactivity was in the form of aggregates; (ii) Herp protein was increased, but its mRNA was not. Accordingly, in s-IBM muscle fibers: (i) increase of Herp might be due to both ER-stress and proteasome inhibition; (ii) co-localization of Herp with Abeta, proteasome, and ER-chaperone GRP78 could reflect its possible role in processing and degradation of cytotoxic proteins in ER.     

蛋白质内含子介导蛛丝功能化平台的建立

为建立高效快捷的蛛丝功能化修饰平台,蛋白质内含子的反式剪接技术被首次应用于重组蛛丝的功能化修饰。在体外通过Ssp Dna B的反式剪接作用,在蛋白质水平上将12 k Da泛素相关修饰蛋白(SUMO)与蛛丝蛋白(W2CT)连接形成功能化蛛丝蛋白SUMOW2CT。修饰后SUMOW2CT与W2CT均能形成纳米至微米级的丝纤维,但SUMOW2CT自动成丝速度明显下降且产量约为W2CT的一半。与W2CT丝纤维(W)相似,SUMOW2CT丝纤维(UW)不具有超收缩能力和对2%SDS不耐受,但机械性能低于W2CT丝纤维。功能化蛋白SUMOW2CT形成的丝纤维中SUMO蛋白仍保持着正确三维结构,可被SUMO蛋白酶酶切。外源功能化蛋白质虽在一定程度上降低了丝的形成速度和机械性能,但修饰上的功能化蛋白仍保持着生物活性,表明断裂蛋白质内含子介导的蛛丝修饰平台成功建立,也为蛛丝的功能化修饰和应用奠定了坚实的技术基础。     

Control of protein adsorption on functionalized electrospun fibers

Electrospun fibers that are protein resistant and functionalized with bioactive signals were produced by solution electrospinning amphiphilic block copolymers. Poly (ethylene glycol)-block-poly(D,L-lactide) (PEG-b-PDLLA) was synthesized in two steps, with a PEG segment of 10 kDa, while the PDLLA block ranged from 20 to 60 kDa. Depending on the PEG and PDLLA segment ratio, as well as solvent selection, the hydrophilicity and protein adsorption could be altered on the electrospun mesh. Furthermore, an alpha-acetal PEG-b-PDLLA was synthesized that allowed the conjugation of active molecules, resulting in surface functionalization of the electrospun fiber. Electrospun material with varying morphologies and diameter were electrospun from 10, 20, and 30 wt.% solutions. Sessile drop measurements showed a reduction in the contact angle from 120 degrees for pure poly(D,L-lactide) with increasing PEG/PDLLA ratio. All electrospun block PEG-b-PDLLA fibers had hydrophilic properties, with contact angles below 45 degrees . The fibers were collected onto six-arm star-poly(ethylene glycol) (star-PEG) coated silicon wafers and incubated with fluorescently labeled proteins. All PEG-b-PDLLA fibers showed no detectable adsorption of bovine serum albumin (BSA) independent of their composition while a dependence between hydrophobic block length was observed for streptavidin adsorption. Fibers of block copolymers with PDLLA blocks smaller than 39 kDa showed no adsorption of BSA or streptavidin, indicating good non-fouling properties. Fibers were surface functionalized with N(epsilon)-(+)-biotinyl-L-lysine (biocytin) or RGD peptide by attaching the molecule to the PEG block during synthesis. Protein adsorption measurements, and the controlled interaction of biocytin with fluorescently labeled streptavidin, showed that the electrospun fibers were both resistant to protein adsorption and are functionalized. Fibroblast adhesion was contrasting between the unfunctionalized and RGD-coupled electrospun fabrics, confirming that the surface of the fibers was functionalized. The PEG-b-PDLLA surface functionalized electrospun fibers are promising substrates for controlling cell-material interactions, particularly for tissue-engineering applications.     

大鼠脊神经节感觉神经特异蛋白35kD(SSP-35)的纯化及鉴定(英文)

 以大鼠脊髓背根神经节及背根纤维组织为材料 ,通过离子交换层析 ,FPLC凝胶过滤等技术分离纯化了脊感觉神经特异蛋白 35kD(SSP 35) .经SDS PAGE分析 ,该蛋白特异地存在于脊感觉神经而不存在于脊运动神经 .非还原电泳结果表明 ,该蛋白分子内含有巯基 ,有二聚体存在 .根据HPLC测定 ,蛋白纯度达 90 %以上 .将此蛋白给予培养的PC 1 2细胞 ,观察到它具有神经营养作用 .