共查询到19条相似文献,搜索用时 93 毫秒
1.
2.
HBV PreS2-MBP融合蛋白在大肠杆菌不同部位的表达 总被引:1,自引:0,他引:1
目的:分析比较HBV PreS2蛋白在大肠杆菌细胞周质和细胞质中的融合表达情况。方法:构建大肠杆菌细胞周质中融合表达HBV PreS2蛋白的载体pMAL-P2x/S2和细胞质中融合表达载体pMAL-C2x/S2,IPTG诱导表达,产物经SDS-PAGE和Western Blot检测,分析HBV PreS2-MBP融合蛋白在菌体不同部位的表达情况及表达产物的存在形式。结果:细胞周质中表达的融合蛋白有一部分被降解了,而细胞质中表达出了完整的融合蛋白。细胞质中表达的融合蛋白以包涵体和可溶性蛋白两种形式存在。结论:HBV PreS2-MBP融合蛋白适合在大肠杆菌细胞质中表达。 相似文献
3.
4.
大肠杆菌周质和外膜蛋白的定位 总被引:1,自引:0,他引:1
大肠杆菌周质和外膜蛋白发挥功能必须首先到达其特定亚细胞分区.大肠杆菌通过一系列与蛋白质分泌有关的蛋白(Sec蛋白)将周质和外膜蛋白转运至内膜.在切除了信号肽后,与周质蛋白的定位不同的是,外膜蛋白的最终定位还需要其他因子的协助.外膜蛋白的定位近来认为是以周质作为中介的. 相似文献
5.
6.
7.
8.
9.
重组蛋白在大肠杆菌中表达时,往往面临着形成包涵体的问题,而重组蛋白若是分泌至周质空间则基本解决了这一问题,周质空间的周质蛋白不仅能帮助重组蛋白正确折叠还有利于二硫键的生成。信号肽是一段由15-30个氨基酸组成,被融合在重组蛋白N端的短肽,按照结构、功能的不同可以划分为N区、H区和C区,具有引导重组蛋白转运至细胞周质空间的作用。本文综述了信号肽的结构组成、作用机理和基本分泌途径,讨论了信号肽的高效转运和筛选方法,总结了在大肠杆菌中重组蛋白融合信号肽实现周质表达的新进展,并对未来高效信号肽选择方面的研究进行了探讨。 相似文献
10.
鲑鱼生长激素基因分泌型表达质粒的构建 总被引:1,自引:0,他引:1
生长激素(GH)是动物垂体前叶分泌的一种多肽类激素.应用分子重组及PCR等技术,构建了一种鲑鱼生长激素基因分泌型表达质粒pOsGH153,使编码鲑鱼生长激素成熟肽的序列克隆在大肠杆菌分泌型表达载体PIN-Ⅲ-ompA内,直接位于编码大肠杆菌外膜蛋白A信号肽序列的下游,在Lpp-Lac杂合启动子控制下,经IPTG诱导,分子量约23 000的鲑鱼生长激素在大肠杆菌中获得高效表达,该产物具有天然鲑鱼生长激素的免疫活性,直接分泌到细胞周质,而信号肽被自动剪除. 相似文献
11.
Cloning of the pectate lyase genes from Erwinia carotovora and their expression in Escherichia coli 总被引:11,自引:0,他引:11
A hybrid cosmid coding for pectate lyase (PL) activity was identified from an Erwinia carotovora genomic library by an immunological screening method. A 7-kb DNA fragment was identified which codes for three proteins identical in size to proteins with PL activity purified from E. carotovora culture supernatants. The three proteins had apparent Mrs of 41, 44 and 44 X 10(3) as estimated by SDS-PAGE. None of the PLs were exported from Escherichia coli strain HB101 but all were found in the periplasmic space. Plant tissue was macerated by the PLs made in E. coli. 相似文献
12.
Co-regulation of the phosphate-binding protein and alkaline phosphatase synthesis in Escherichia coli. 总被引:6,自引:5,他引:1
下载免费PDF全文
![点击此处可从《Journal of bacteriology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
In phosphate-starved cells of Escherichia coli, the synthesis of alkaline phosphatase and some additional periplasmic proteins is derepressed. One of these proteins, which does not appear in a phoS- constitutive strain, has been identified as well the periplasmic phosphate-binding protein. 相似文献
13.
Periplasmic protein associated with the oligopeptide permeases of Salmonella typhimurium and Escherichia coli. 总被引:14,自引:10,他引:4
A periplasmic protein essential for the function of the oligopeptide transport system of Salmonella typhimurium was identified. This protein, encoded by the oppA gene, is one of the most abundant proteins in the periplasm and, with an apparent molecular weight of 52,000, is considerably larger than any other known periplasmic transport component. A similarly abundant periplasmic protein forms part of the oligopeptide transport system of Escherichia coli. 相似文献
14.
Many proteins require enzymatic assistance in order to achieve a functional conformation. One rate-limiting step in protein folding is the cis-trans isomerization of prolyl residues, a reaction catalyzed by prolyl isomerases. SurA, a periplasmic protein of Escherichia coli, has sequence similarity with the prolyl isomerase parvulin. We tested whether SurA was involved in folding periplasmic and outer membrane proteins by using trypsin sensitivity as an assay for protein conformation. We determined that the efficient folding of three outer membrane proteins (OmpA, OmpF, and LamB) requires SurA in vivo, while the folding of four periplasmic proteins was independent of SurA. We conclude that SurA assists in the folding of certain secreted proteins. 相似文献
15.
Using biotinylated phage (BIO-phages), we observed the infection of filamentous phages into Escherichia coli JM109 morphologically. BIO-phages and BIO-phage-derived proteins, mainly pVIII, were detected in E. coli by using the avidin-biotin-peroxidase complex method with electron microscopy. Infected cells revealed positive staining on the outer and inner membranes and in the periplasmic space. Some cells showed specific or predominant staining of the outer membrane, whereas others showed predominant staining of the inner membrane or equivalent staining of the outer and inner membranes. The periplasmic spaces in some infected cells were expanded and filled with reaction products. Some cells showed wavy lines of positive staining in the periplasmic space. BIO-phages were detected as thick filaments or clusters covered with reaction products. The ends of the infecting phages were located on the surface of cells, in the periplasmic space, or on the inner membrane. These findings suggest that phage major coat proteins are integrated into the outer membrane and that phages cause periplasmic expansion during infection. 相似文献
16.
Characterization of an Escherichia coli rotA mutant, affected in periplasmic peptidyl-prolyl cis/trans isomerase 总被引:2,自引:1,他引:1
The rotA gene of Escherichia coli encodes a peptidyl-prolyl cis/trans isomerase (PPlase), which is supposed to catalyse protein folding in the periplasm. To investigate the importance of the enzyme, the rotA gene was cloned and a chromosomal deletion mutant was created. The rotA mutant was normally viable. No residual PPlase activity could be detected in the periplasmic fraction of the mutant. Comparison of the patterns of periplasmic and outer membrane proteins by SDS-PAGE revealed no differences in protein composition between the rotA mutant and its parental strain. Similarly, the kinetics of periplasmic protein folding and outer membrane protein assembly appeared unaffected by the rotA mutation. Our results show that the periplasmic PPlase of E. coli is not essential and that the protein does not play an important role in protein folding. 相似文献
17.
J M Brass C F Higgins M Foley P A Rugman J Birmingham P B Garland 《Journal of bacteriology》1986,165(3):787-795
We have introduced biologically active, fluorescently labeled maltose-binding protein into the periplasmic space of Escherichia coli and measured its lateral diffusion coefficient by the fluorescence photobleaching recovery method. Diffusion of this protein in the periplasm was found to be surprisingly low (lateral diffusion coefficient, 0.9 X 10(-10) cm2 s-1), about 1,000-fold lower than would be expected for diffusion in aqueous medium and almost 100-fold lower than for an equivalent-size protein in the cytoplasm. Galactose-binding protein, myoglobin, and cytochrome c were also introduced into the periplasm and had diffusion coefficients identical to that determined for the maltose-binding protein. For all proteins nearly 100% recovery of fluorescence was obtained after photobleaching, indicating that the periplasm is a single contiguous compartment surrounding the cell. These data have considerable implications for periplasmic structure and for the role of periplasmic proteins in transport and chemotaxis. 相似文献
18.
19.
A Shewanella expression system has been used for an overproduction of c-type multiheme proteins. The proteins were exported to the periplasmic space for the maturation. Since the periplasmic expression system is attractive, especially for protease-sensitive proteins, an expression vector containing a signal peptide was constructed for expressions in the periplasmic space of Shewanella oneidensis. To evaluate the system, two eukaryotic proteins which originally do not have signal sequences and are difficult to express in Escherichia coli, were selected. The first is human cytochrome c. Properties of the recombinant cytochrome c were identical to those previously reported, indicating the protein is intact. The other was potato calcium-dependent protein kinase. The protein was expressed in periplasmic space. These results indicated that the system is generally applicable for any protein expression including c-type cytochromes, protease-sensitive proteins and those with multi-disulfide bonds because of transportation to the periplasmic space. 相似文献