• BMB Reports
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• v.52 no.7
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• pp.424-433
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• 2019

Autism spectrum disorder (ASD) is a complex neurodevelopmental monogenic disorder with a strong genetic influence. Idiopathic autism could be defined as a type of autism that does not have a specific causative agent. Among signalling cascades, mTOR signalling pathway plays a pivotal role not only in cell cycle, but also in protein synthesis and regulation of brain homeostasis in ASD patients. The present review highlights, underlying mechanism of mTOR and its role in altered signalling cascades as a triggering factor in the onset of idiopathic autism. Further, this review discusses how distorted mTOR signalling pathway stimulates truncated translation in neuronal cells and leads to downregulation of protein synthesis at dendritic spines of the brain. This review concludes by suggesting downstream regulators such as p70S6K, eIF4B, eIF4E of mTOR signalling pathway as promising therapeutic targets for idiopathic autistic individuals.

• Molecules and Cells
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• v.26 no.2
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• pp.140-145
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• 2008

Expression of olive flounder hepcidin I (HepI) fused with truncated OmpA signal peptides ($OmpASP_{tr}$) as directional signals does not produce soluble fusion proteins. However, by inserting amino acid segments (xxx) varying in pI and hydrophobicity/hydrophilicity into a leader sequence containing a truncated OmpASP ($OmpASP_{tr}$) and a factor Xa cleavage site (Xa) [$OmpASP_{tr}{\mid}(xxx){\mid}Xa$], we were able in some cases to express soluble recombinant HepI. Soluble expression of the recombinant protein strongly correlated with (xxx) insertions of high pI and hydrophilicity. Therefore, we modified the $OmpASP_{tr}{\mid}(xxx){\mid}Xa$ sequence by inserting Arg and Lys into (xxx) to increase the hydrophilicity of the signal peptide region. These modifications enhanced the expression of soluble recombinant HepI. Hydropathic profile analysis of the $OmpASP_{tr}{\mid}(xxx){\mid}Xa$ HepI fusion proteins revealed that the transmembrane-like domains derived from the $OmpASP_{tr}{\mid}(xxx){\mid}Xa$ sequence were larger than the internal positively charged domain native to HepI. It should therefore be possible to overcome the obstacle of internal positively charged domains to obtain soluble expression of recombinant proteins by monitoring the hydrophilicity and hydropathic profile of the signal peptide region using a computer program.

• The Korean Journal of Physiology and Pharmacology
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• v.21 no.6
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• pp.667-674
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• 2017

Angiotensin II (Ang II) is metabolized from N-terminal by aminopeptidases and from C-terminal by Ang converting enzyme (ACE) to generate several truncated angiotensin peptides (Angs). The truncated Angs have different biological effects but it remains unknown whether Ang-(4-8) is an active peptide. The present study was to investigate the effects of Ang-(4-8) on hemodynamics and atrial natriuretic peptide (ANP) secretion using isolated beating rat atria. Atrial stretch caused increases in atrial contractility by 60% and in ANP secretion by 70%. Ang-(4-8) (0.01, 0.1, and $1{\mu}M$) suppressed high stretch-induced ANP secretion in a dose-dependent manner. Ang-(4-8) ($0.1{\mu}M$)-induced suppression of ANP secretion was attenuated by the pretreatment with an antagonist of Ang type 1 receptor ($AT_1R$) but not by an antagonist of $AT_2R$ or $AT_4R$. Ang-(4-8)-induced suppression of ANP secretion was attenuated by the pretreatment with inhibitor of phospholipase (PLC), inositol triphosphate ($IP_3$) receptor, or nonspecific protein kinase C (PKC). The potency of Ang-(4-8) to inhibit ANP secretion was similar to Ang II. However, Ang-(4-8) $10{\mu}M$ caused an increased mean arterial pressure which was similar to that by 1 nM Ang II. Therefore, we suggest that Ang-(4-8) suppresses high stretch-induced ANP secretion through the $AT_1R$ and $PLC/IP_3/PKC$ pathway. Ang-(4-8) is a biologically active peptide which functions as an inhibition mechanism of ANP secretion and an increment of blood pressure.

• Journal of Microbiology and Biotechnology
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• v.28 no.12
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• pp.2036-2045
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• 2018

An endo-${\beta}$-1,4-glucanase gene, cel5L, was cloned using the shot-gun method from Bacillus sp.. The gene, which contained a predicted signal peptide, encoded a protein of 496 amino acid residues, and the molecular mass of the mature Cel5L was estimated to be 51.8 kDa. Cel5L contained a catalytic domain of glycoside hydrolase (GH) family 5 and a carbohydrate-binding module family 3 (CBM_3). Chromatography using HiTrap Q and CHT-II resulted in the isolation of two truncated forms corresponding to 50 (Cel5L-p50) and 35 kDa (Cel5L-p35, CBM_3-deleted form). Both enzymes were optimally active at pH 4.5 and $55^{\circ}C$, but had different half-lives of 4.0 and 22.8 min, respectively, at $70^{\circ}C$. The relative activities of Cel5L-p50 and Cel5L-p35 for barley ${\beta}$-glucan were 377.0 and 246.7%, respectively, compared to those for carboxymethyl-cellulose. The affinity and hydrolysis rate of pNPC by Cel5L-p35 were 1.7 and 3.3 times higher, respectively, than those by Cel5L-p50. Additions of each to a commercial enzyme set increased saccharification of pretreated rice straw powder by 17.5 and 21.0%, respectively. These results suggest CBM_3 is significantly contributing to thermostability, and to affinity and substrate specificity for small substrates, and that these two enzymes could be used as additives to enhance enzymatic saccharification.

• BMB Reports
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• v.34 no.3
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• pp.201-205
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• 2001

Thionins are a family of low molecular weight cysteine-rich antimicrobial peptides. We isolated a cDNA encoding thionin gene from a flower bud cDNA library of Chinese cabbage (CFT). The gene contains 611 by nucleotides with 60 bp, and 150 by untranslated regions at its N- and C-terminal, respectively. The deduced amino acid sequence encoded 133 amino acids containing precursor polypeptide. The protein reveals that the precursor has a tripartite structure: a putative signal sequence at the N-terminus, followed by a mature thionin peptide, and a C-terminal acidic domain, which facilitates transport of the mature thionin through membrane. Genomic Southern blot analysis suggests that the CFT gene may be present as a single or two copy gene in the Chinese cabbage genome. Northern blot analysis shows that the gene is specifically expressed in flowers, but not in leaves, stems, or roots. When we analyzed the antifungal activity of the recombinant CFT protein, which was expressed in E. coli using the truncated cDNA region corresponding to the mature protein part, it was not active on fungal growth inhibition.

• Journal of Microbiology and Biotechnology
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• v.30 no.11
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• pp.1777-1784
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• 2020

To increase the availability of microalgae as producers of valuable compounds, it is necessary to develop novel systems for gene expression regulation. Among the diverse expression systems available in microalgae, none are designed to induce expression by low temperature. In this study, we explored a cold-inducible system using the antifreeze protein (AFP) promoter from a polar diatom, Chaetoceros neogracile. A vector containing the CnAFP promoter (pCnAFP) was generated to regulate nuclear gene expression, and reporter genes (Gaussia luciferase (GLuc) and mVenus fluorescent protein (mVenus)) were successfully expressed in the model microalga, Chlamydomonas reinhardtii. In particular, under the control of pCnAFP, the expression of these genes was increased at low temperature, unlike pAR1, a promoter that is widely used for gene expression in C. reinhardtii. Promoter truncation assays showed that cold inducibility was still present even when pCnAFP was shortened to 600 bp, indicating the presence of a low-temperature response element between -600 and -477 bp. Our results show the availability of new heterologous gene expression systems with cold-inducible promoters and the possibility to find novel low-temperature response factors in microalgae. Through further improvement, this cold-inducible promoter could be used to develop more efficient expression tools.

• Journal of Microbiology and Biotechnology
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• v.6 no.3
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• pp.219-220
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• 1996

The pKH2 isolated from the multidrug-resistant Staphylococcus aureus SA2 is a 40.98-kb plasmid and mediates resistance to ampicillin, clindamycin, erythromycin, kanamycin, and streptomycin. The 3.4-kb HindIII fragment conferring kanamycin resistance was cloned from the pKH2 into pBluescriptII $KS^+$ and partial sequence determination of that fragment was carried out. Sequence analysis revealed that the kanamycin resistance gene which encoded aminoglycoside 3'-phosphotransferase was linked to the streptothricin resistance gene. But a nonsense mutation was found in the streptothricin resistance gene and this mutation resulted in a truncated protein of streptothricin acetyltransferase. Homology comparison with nucleotide sequence databases revealed that the 3.4-kb HindIII fragment of pKH2 had been derived not from S. aureus but from Gram-negative Campylobacter coli.

• BMB Reports
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• v.51 no.6
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• pp.263-264
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• 2018

We identified osteoclast-derived SLIT3 as a new coupling factor using fractionated secretomics. Coupling links bone resorption to bone formation. SLIT3 stimulated the recruitment and proliferation of osteoblasts into bone remodeling sites via activation of ${\beta}-catenin$. Autocrine signaling by SLIT3 also inhibited bone resorption by suppressing the fusion and differentiation of pre-osteoclasts. All mice lacking Slit3 or its receptor Robo1 showed an osteopenic phenotype with low bone formation and high bone resorption. A small truncated recombinant SLIT3 protein increased bone mass in an osteopenic mouse model. These results suggest that SLIT3 is a novel therapeutic target in metabolic bone diseases.

• Applied Biological Chemistry
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• v.41 no.1
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• pp.23-30
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• 1998

The cryIB, truncated cryIB$[cryIB({\alpha})]$, cryIA(b), and cytA genes, encoding 135-, 89-, 131-, and 27-kDa proteins, respectively, from Bacillus thuringiensis were cloned into a shuttle vector pBES and expressed in E. coli and Bacillus species. The morphology and solubility in alkaline buffer of the insecticidal crystal proteins were investigated. Transformation of intact cells of E. coli and Bacillus species was achieved by electroporation. High field strength of 11.0 kV/cm and resistance of 129 ohms were required for efficient transformation of E. coli strains and 4.5 kV/cm and 48 ohms for Bacillus species. Strains of recombinant E. coli and Bacillus species produced the insecticidal crystal proteins and accumulated as the same bipyramidal and irregular structures as those of CryIB and IA(b) and CytA of B. thuringiensls, respectively. The insecticidal crystal proteins accumulated in recombinant E. coli wire smaller in size than those in recombinant Bacillus species. The solubility in alkaline buffer of the insecticidal crystal proteins of recombinant E. coli increased gradually as the pH increased, whereas in the case of Bacillus species the solubility increased gradually as the pH increased up to 9 and then the solubility increased greatly up to two times higher than that of E. coli proteins.

• Journal of Gastric Cancer
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• v.5 no.4 s.20
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• pp.260-265
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• 2005

Purpose: The protein kinase Chk1 is required for cell cycle arrest in response to DNA damage and is shown to play an important role in the G2/M checkpoint. The aim of this study was to investigate the relationship between microsatellite instability and frameshift mutation of the Chk1 gene in gastric cancers. Materials and Methods: The microsatellite instability was analyzed in 95 primary gastric carcinomas by using microdissection and 6 microsatellite markers. We also peformed single strand conformational polymorphism and sequencing to detect frameshift mutation of the Chk1 gene. Results: We found positive microsatellite instability in 19 (20%) of the 95 gastric cancers, 13 high- and 6 low-frequency microsatellite instability cases. The frameshift mutation of Chk1, which resulted in a truncated Chk1 protein, was detected in two high-frequency microsatellite instability cases. Conclusion: These data suggest that the microsatellite instability may contribute to the development of gastric carcinomas through inactivation of Chk1.