• Journal of Microbiology and Biotechnology
• /
• v.18 no.2
• /
• pp.228-233
• /
• 2008

Effects of chaperones on mRNA stability and gene expression were studied in order to develop an efficient Escherichia coli expression system that can maximize gene expression. The stability of mRNA was modulated by introducing various secondary structures at the 5'-end of mRNA. Four vector systems providing different 5'-end structures were constructed, and genes encoding GFPuv and endoxylanase were cloned into the four vector systems. Primer extension assay revealed different mRNA half-lives depending on the 5'-end secondary structures of mRNA. In addition to the stem-loop structure at the 5'-end of mRNA, coexpression of dnaK-dnaJ-grpE or groEL-groES, representative heat-shock genes in E. coli, increased the mRNA stability and the level of gene expression further, even though the degree of stabilization was varied. Our work suggests that some of the heat-shock proteins can function as mRNA stabilizers as well s protein chaperones.

• Biotechnology and Bioprocess Engineering:BBE
• /
• v.11 no.5
• /
• pp.414-419
• /
• 2006

When the alginate lyase gene (aly) from Pseudoalteromonas elyakovii was expressed in E. coli, most of the gene product was organized as aggregated insoluble particles known as inclusion bodies. To examine the effects of chaperones on soluble and nonaggregated form of alginate lyase in E. coli, we constructed plasm ids designed to permit the coexpression of aly and the DnaK/DnaJ/GrpE or GroEL/ES chaperones. The results indicate that coexpression of aly with the DnaK/DnaJ/GrpE chaperone together had a marked effect on the yield alginate lyase as a soluble and active form of the enzyme. It is speculated this result occurs through facilitation of the correct folding of the protein. The optimal concentration of L-arabinose required for the induction of the DnaK/DnaJ/GrpE chaperone was found to be 0.05mg/mL. An analysis of the protein bands on SDS-PAGE gel indicated that at least 37% of total alginate lyase was produced in the soluble fraction when the DnaK/DnaJ/GrpE chaperone was coexpressed.

• Proceedings of the Korean Society of Sericultural Science Conference
• /
• 2001.10a
• /
• pp.30-32
• /
• 2001

no Abstract, See Full Text

• BMB Reports
• /
• v.42 no.10
• /
• pp.623-630
• /
• 2009

Hsp90, an evolutionarily conserved molecular chaperone, is involved in the folding, stabilization, activation, and assembly of a wide range of 'client' proteins, thus playing a central role in many biological processes. Especially, several oncoproteins act as Hsp90 client proteins and tumor cells require higher Hsp90 activity than normal cells to maintain their malignancy. For this reason, Hsp90 has emerged as a promising target for anti-cancer drug development. It is still largely unknown how Hsp90 can recognize structurally unrelated client proteins. However, recent progress in structural studies on Hsp90 and its interaction with various co-chaperones has broadened our knowledge of how the Hsp90 ATPase activity, which is essential for its chaperone function, is regulated and coupled with the conformational changes of Hsp90 dimer. This review focuses on the roles of various Hsp90 co-chaperones in the regulation of the Hsp90 ATPase cycle, as well as in the selection of client proteins. In addition, the current development of Hsp90 inhibitors based on the structural information will be discussed.

• Biomedical Science Letters
• /
• v.14 no.1
• /
• pp.59-62
• /
• 2008

This experiment was performed to study the effect of TSH (thyroid-stimulating hormone) on the expression of endoplasmic reticulum (ER) chaperones in the rat thyrocytes FRTL-5 cells. Although the expressions of ER membrane kinases (ATF6, IRE1 and PERK) were specially enhanced under absence of TSH, no remarkable up- or down regulations of ER chaperones (BiP, CHOP and Calnexin) were detected by TSH. We firstly report here that TSH by dose up-regulated expression of ER membrane kinases in FRTL-5 culture thyrocytes.

• Mycobiology
• /
• v.43 no.3
• /
• pp.272-279
• /
• 2015

To screen molecular chaperones similar to small heat shock proteins (sHsps), but without ${\alpha}$-crystalline domain, heat-stable proteins from Schizosaccharomyces pombe were analyzed by 2-dimensional electrophoresis and matrix assisted laser desorption/ionization time-of-flight mass spectrometry. Sixteen proteins were identified, and four recombinant proteins, including cofilin, NTF2, pyridoxin biosynthesis protein (Snz1) and Wos2 that has an ${\alpha}$-crystalline domain, were purified. Among these proteins, only Snz1 showed the anti-aggregation activity against thermal denaturation of citrate synthase. However, pre-heating of NTF2 and Wos2 at $70^{\circ}C$ for 30 min, efficiently prevented thermal aggregation of citrate synthase. These results indicate that Snz1 and NTF2 possess molecular chaperone activity similar to sHsps, even though there is no ${\alpha}$-crystalline domain in their sequences.

• Journal of Life Science
• /
• v.23 no.9
• /
• pp.1104-1108
• /
• 2013

Ethylene glycol (EG) is the most commonly used for automotive antifreeze, and it's easily misuseful for human. EG poisoning occurs in suicide attempts and infrequently, either intentionally through misuse or accidentally because of sweet taste. Though EG itself is mild toxic to the human body, it becomes higher toxic organic acids by in vivo broken down that are responsible for extensive cellular damage in various tissues caused principally by the metabolites. It is already well known that various cellular stresses induce gene expression of endoplasmic reticulum (ER) chaperones and ER stress sensors. This study demonstrated that regulation of gene expression of ER chaperones and ER stress sensors was induced by EG in rat tissues, and in tissues histological changes are also detected by both staining H&E and immunofluorescent.

• Microbiology and Biotechnology Letters
• /
• v.43 no.3
• /
• pp.212-218
• /
• 2015

Fucosyltransferases (FucTs) catalyze fucosyl transfer from guanosine-diphosphate fucose (GDP-β-L-fucose) to acceptor molecules to form fucosyloligosaccharides with α-glycosidic linkages. However, when FucT genes have been expressed in Escherichia coli, most cases have resulted in the production of inclusion bodies. In this study, to overcome this drawback, molecular chaperones were co-expressed with α1,2-fucosyltransferase (FucT2) in E. coli. For this, the pACYC184 vector, having genes for chaperones such as GroEL, GroES, DnaK, DnaJ, and GrpE, were transformed into E. coli BL21 (DE3) star harboring pHFucT2, including the FucT2 gene from Helicobacter pylori 26695. The results from SDS-PAGE showed that 5 chaperones were successfully expressed and the soluble fraction of FucT2 was also increased. HPLC analysis revealed that the coexpression of chaperone proteins resulted in a 5-fold increase in the total activity of fucosyltransferase in E. coli. In conclusion, the FucT2 expression system developed in this study can be used as a useful tool for the synthesis of fucosyloligosaccharides.

• Journal of Physiology & Pathology in Korean Medicine
• /
• v.16 no.4
• /
• pp.680-683
• /
• 2002

Bovine aortic endothelial cells(BAEC) produce two variant forms of laminin with a subunit composition of AB1B2 and A'B1B2. Analyses of the intracellular assembly of these subunits revealed that the B1B2 dimer formed first, and that A or A' joined to form the AB1B2 or A'B1B2 trimer. Angiostatic steroids shifted the relative size of the A and A' monomer pool in BAEC, and competition between the A and A' subunits in joining the B1B2 dimer produced AB1B2 and A'B1B2 in different ratios. This result suggests that subunit replacement is the general mechanism for producing laminin variants by various cells for tissue morphogenesis. When laminin subunits in BAEC were cross-linked with dithio-bis-succinimidylpropionate(DSP) and immunoprecipitated with anti-Iaminin antiserum, monomeric A,A',B1 and B2 monomers and the B1B2 dimer migrated as extremely large molecules in sodium dodecyl sulfate gel electrophoresis under nonreducing conditions. When the crosslinking disulfide bonds were cleaved under reducing conditions, they migrated as the usual subunits. This result suggests that molecular chaperones were involved in the process of the assembly and replacement of laminin subunits.

• Journal of Life Science
• /
• v.12 no.2
• /
• pp.53-56
• /
• 2002

The baculovirus expression vector system (BEVS) is one of the powerful heterologous protein expression systems using insect cells. As a result this has become a hot issue in the fleld of biotechnology. The advantage of the BEVS is that the large-scale production of heterologous proteins, which undergo posttranslational modification in the endoplasmic reticulum (ER), can be accomplished. Altrough posttranslational modification of heterologous proteins in insect cells is more similar to mammalian cells than yeast, it is not always identical. Therefore, aggregation and degradation can sometimes occur in the ER. To produce a high level of bioactive heterologous proteins using BEVS in insect cells, the prerequisite is to completely understand the posttranslational conditions that determine how newly synthesized polypeptides are folded and assembling with ER chaperones in the ER lumen. Here, we provide information on current BEVS problems and the possibility of successful heterologous protein production from mammalian cells.