• Development and Reproduction
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• v.26 no.4
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• pp.165-174
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• 2022

Three proteins [myosin heavy chain (MHC), filamin-C fragment (FIL-C), and actin 2 (ACT2)] were identified in adductor muscle from diploid and triploid Pacific oysters (Crassostrea gigas) and the relationship between the condition index (CI) and mRNA expression of these genes was investigated, together with the mRNA expression of molluscan insulin-related peptide (MIP), C. gigas insulin receptor-related receptor (CIR), and insulin-like growth factor binding protein complex acid labile subunit (IGFBP-ALS). Monthly changes in the CI were similar to the changes in the tissue weight rate in both groups. ACT2 and MHC mRNA expression was statistically higher in the triploid than the diploid, while FIL-C mRNA expression was significantly higher in the diploid (p<0.05). The MIP, CIR, and IGFBP-ALS mRNA expression of the diploid oysters were all significantly higher in July than in other months (p<0.05). The MIP, CIR, and IGFBP-ALS mRNA expression in the triploid oysters was high in July, but there were no significant differences (p>0.05). Changes in the expression levels of the genes investigated in this study could be used as intrinsic indicators of the annual growth, maturity, and spawning period of cultured diploid and triploid C. gigas in Tongyeong, Korea.

• Proceedings of the Korea Society of Poultry Science Conference
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• 2005.11a
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• pp.64-65
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• 2005

Members of the Pumilio are the RNA binding proteins acting as translational repressors and required for germ cell development and asymmetric division. We identified chicken Pum1 and Pum2 that are similar to mouse and human in highly conserved C-terminal RNA-binding domain and eight tandem repeats. The comparative sequence analysis of Pum1 and Pum2 from fly, chicken, mouse and human shows high degree of evolutionary conservation in the homology of the peptide sequence and the structure of PUM-HD (Pumilio homology domain) with similar spacing between adjacent Pum repeats. Also, structures of chicken Pum1 and Pum2 genes are almost identical to those of mouse and human. We revealed that the expression levels of Pum1 and Pum2 were the highest in hatched female gonad among various embryonic tissues, and Pum2 expressed highly in 12-day and hatched gonad by real-time RT-PCR. These results suggest that Pum1 and Pum2 might have an effect on the development of chicken gonad.

• Journal of Life Science
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• v.18 no.7
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• pp.931-939
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• 2008

The purpose of this study was to determine the effect of energy supplement on responses of plasma insulin-like growth factor (IGF)-1 and IGF binding proteins (IGFBPs) to growth hormone-releasing peptide-2 (GHRP-2) administration in normal protein-fed wethers, and to observe the effect of GHRP-2 treatment on hepatic growth hormone (GH) receptor in well-fed wethers. Plasma IGF-1 and 39-42 kDa IGFBP-3 during the HENP (CP, crude protein 0.34 and TDN, total digestible nutrients 1.83 kg/day DM, dry matter intake) treatment period were higher than in the LENP (CP 0.32 kg and TDN 0.87 kg/day DM intake) period (P<0.05). The response of GH was stimulated by GHRP-2 ($12.5\;{\mu}g/kg$ body weight/day) administration during both of the feed treatment periods (P<0.05). The area under curve (AUC) increment and average concentration of GH (0-180 min) with GHRP-2 administration was higher during HENP treatment than LENP treatment (P<0.01). During the HENP treatment period from day 1 to day 7 of twice daily GHRP-2 treatment, the plasma IGF-1 increment was increased on days 2, 6 and 7 of GHRP-2 administration (P<0.05). On the basis of ligand blotting, the proportions of plasma 39-43 kDa IGFBP-3 during the HENP treatment period only showed a significant difference on days 6 and 7 with GHRP-2 administration. No significant difference in the specific binding of $^{125}I-labeled$ oGH to hepatic membranes was detected between the saline and GHRP-2 treatments of the HENP-fed wethers. These results suggest that the nutritional balance between energy and protein may affect the endogenous GH / IGF-1 axis as well as plasma IGFBP-3 levels.

• The Journal of the Korean Society for Microbiology
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• v.12 no.1
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• pp.19-32
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• 1977

The specificity of the N- and C-terminal antigenic determinant($P_{17}$: sequence $Lys^1-{cys-}^6-Asn^{27},\;{Trp^{12}}_2-Cys^{127}-Leu^{129}$) of hen egg-white lysozyme(HL) was studied in more detail. In a Scatchard plot of the binding of $^{14}C$-acetyl HL with guinea pig purified anti-$P_{17}$ antibody experimental values bent sharply aear r=1. This suggests of two antibody populations with different affinities for HL or possible steric hindrance in the binding of a second HL molecule to the second binding site of the antibody molecule. The antigenic activities of various peptides were tested by measuring their inhibition of the binding of $^{14}C-acetyl-P_{17}$ with the antibody, Only $P_{17}$ and $P_{17}t$(sequence $Lys^1-cys^6-Homoser^{12},\;Trp^{123}-Cys^{127}-Leu^{128})$) were inhibitory, with $K_1$ values of $2.0{\times}10^4$ and $8.1{\times}10^3$, respectively. These results indicate that the direct binding site of $P_{17}$ to anti-$P_{17}$ antibody may be located in the terminal portion of $P_{17}$ (sequence $Lys^1-Cys^6-Homoser^{12},\;Trp^{123}-Cys^{127}-Leu^{129})$) while the rest of $P_{17}$ may be important in maintaining the conformation of this determinant. The single disulphide bond involved in this determinant is essential for manifestation of immunological activity.

• Journal of Applied Biological Chemistry
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• v.59 no.3
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• pp.221-225
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• 2016

Tolaasin is a pore-forming peptide toxin produced by Pseudomonas tolaasii and causes a brown blotch disease by disrupting membrane structures of cultivated mushrooms. The mechanism and characteristics of tolaasin pore formation are not known in detail; however, tolaasin pores have been demonstrated in the artificial lipid bilayer. Since the tolaasin pore appeared less frequently and unstable in lipid bilayer, a mismatch between the length of tolaasin pore and the thickness of lipid membrane had been suggested. Therefore, tolaasin-induced hemolyses were measured by the additions of phospholipids composed of various fatty acids with different carbon numbers. When phosphatidylethanolamines made with two decanoic acids (C10:0, 1,2-didecanoyl-sn-glycero-3-phosphoethanolamine; DDPE), myristic acids (C14:0, 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine), and stearic acids (C18:0, 1,2-distearoyl-sn-glycero-3-phosphoethanolamine) were added to the buffer containing RBCs and tolaasin peptides, DDPE facilitated the tolaasin-induced hemolysis while the other two phospholipids showed no effects. At various concentrations of DDPE, the tolaasin-induced hemolysis was stimulated as a dose-dependent manner. The phospholipids composed of mediumchain fatty acids stabilize the tolaasin pore probably by binding between the pore structure and membrane phospholipids and making the membrane thickness thinner around the pore. These results showed that tolaasin molecules make more stable pores in the membrane made with phospholipids composed of medium length fatty acids, suggesting that the length of tolaasin pore is a little shorter than the thickness of RBC membrane.

• Microbiology and Biotechnology Letters
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• v.45 no.2
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• pp.155-161
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• 2017

A gene coding for the xylanase predicted from the partial genomic sequence of Paenibacillus woosongensis was cloned by PCR amplification and sequenced completely. This xylanase gene, designated xyn11B, consisted of 1,071 nucleotides encoding a polypeptide of 356 amino acid residues. Based on the deduced amino acid sequence, Xyn11B was identified to be a modular enzyme, including a single carbohydrate-binding module besides the catalytic domain, and was highly homologous to xylanases belonging to glycosyl hydrolase family 11. The SignalP4.1 server predicted a stretch of 26 residues in the N-terminus to be the signal peptide. Using DEAE-Sepharose and Phenyl-Sepharose column chromatography, Xyn11B was partially purified from the cell-free extract of recombinant Escherichia coli carrying a copy of the P. woosongensis xyn11B gene. The partially purified Xyn11B protein showed maximal activity at $50^{\circ}C$ and pH 6.5. The enzyme was more active on arabinoxylan than on oat spelt xylan and birchwood xylan, whereas it did not exhibit activity towards carboxymethylcellulose, mannan, and para-nitrophenyl-${\beta}$-xylopyranoside. The activity of Xyn11B was slightly increased by $Ca^{2+}$ and $Mg^{2+}$, but was significantly inhibited by $Cu^{2+}$, $Ni^{2+}$, $Fe^{3+}$, and $Mn^{2+}$, and completely inhibited by SDS.

• Korean Journal of Microbiology
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• v.52 no.3
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• pp.336-343
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• 2016

A mannanase gene was cloned into Escherichia coli from Cellulosimicrobium sp. YB-43, which had been found to produce two kinds of mannanase, and sequenced completely. This mannanase gene, designated manB, consisted of 1,284 nucleotides encoding a polypeptide of 427 amino acid residues. Based on the deduced amino acid sequence, the ManB was identified to be a modular enzyme including two carbohydrate binding domains besides the catalytic domain, which was highly homologous to mannanases belonging to the glycosyl hydrolase family 5. The N-terminal amino acid sequence of ManB, purified from a cell-free extract of the recombinant E. coli carrying a Cellulosimicrobium sp. YB-43 manB gene, has been determined as QGASAASDG, which was correctly corresponding to signal peptide predicted by SignalP4.1 server for Gram-negative bacteria. The purified ManB had a pH optimum for its activity at pH 6.5~7.0 and a temperature optimum at $55^{\circ}C$. The enzyme was active on locust bean gum (LBG), konjac and guar gum, while it did not exhibit activity towards carboxymethylcellulose, xylan, starch, and para-nitrophenyl-${\beta}$-mannopyranoside. The activity of enzyme was inhibited very slightly by $Mg^{2+}$, $K^+$, and $Na^+$, and significantly inhibited by $Cu^{2+}$, $Zn^{2+}$, $Mn^{2+}$, and SDS. The enzyme could hydrolyze mannooligosaccharides larger than mannobiose, which was the most predominant product resulting from the ManB hydrolysis for mannooligosaccharides and LBG.

• Bulletin of the Korean Chemical Society
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• v.31 no.6
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• pp.1519-1526
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• 2010

The behavior of peptide or protein solutes in saline aqueous solution is a fundamental topic in physical chemistry. Addition of ions can strongly alter the thermodynamic and physical properties of peptide molecules in solution. In order to study the effects of added ionic salts on protein conformation and dynamics, we have used the molecular dynamics (MD) simulations to investigate the behavior of Staphylococcus aureus Hfq protein under two different ionic concentrations: 0.1 M NaCl and 1.0 M NaCl in presence and absence of RNA (a hepta-oligoribonucleotide AU5G). Hfq, a global regulator of gene expression is highly conserved and abundant RNA-binding protein. It is already reported that in vivo the increase of ionic strength results in a drastic reduction of Hfq affinity for $Q{\beta}$ RNA and reduces the tendency of aggregation of Escherichia coli host factor hexamers. Our results revealed the crucial role of 0.1 M NaCl Hfq system on the bases with strong hydrogen bonding interactions and by stabilizing the aromatic stacking of Tyr42 residue of the adjacent subunits/monomers with the adenine and uridine nucleobases. An increase in RNA pore diameter and weakened compactness of the Hfq-RNA complex was clearly observed in 1.0 M NaCl Hfq system with bound RNA. Aggregation of monomers in Hfq and the interaction of Hfq with RNA are greatly affected due to the presence of high ionic strength. Higher the ionic concentration, weaker is the aggregation and interaction. Our results were compatible with the experimental data and this is the first theoretical report for the experimental study done in 1980 by Uhlenbeck group for the present system.

• Korean Journal of Microbiology
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• v.54 no.2
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• pp.126-135
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• 2018

The characteristics of enzyme and gene for mannanase B had been reported from Cellulosimicrobium sp. YB-43 producing some kind of mannanase. A gene coding for the enzyme, named mannanase C (ManC), was expected to be located downstream of the manB gene. The manC gene was cloned by polymerase chain reaction and sequenced completely. From this nucleotide sequence, ManC was identified to consist of 448 amino residues and contain a carbohydrate binding domain CBM2 besides a catalytic domain, which was homologous to mannanase belonging to the glycosyl hydrolase family 5. The catalytic domain of ManC showed the highest amino acid sequence similarity of 55% with the mannanases from Streptomyces sp. SirexAA-E (55.8%; 4FK9_A) and S. thermoluteus (57.6%; BAM62868). The His-tagged ManC (HtManC) lacking N-terminal signal peptide with hexahistidine at C-terminus was produced and purified from cell extract of recombinant Escherichia coli. The purified HtManC showed maximal activity at $65^{\circ}C$ and pH 7.5, with no significant change in its activity at pH range from 7.5 to 10. HtManC showed more active on konjac and locust bean gum (LBG) than guar gum and ivory nut mannan (ivory nut). Vmax and Km values of the HtManC for LBG were 68 U/mg and 0.45 mg/ml on the optimal condition, respectively. Mannobiose and mannotriose were observed on TLC as major products resulting from the HtManC hydrolysis of mannooligosacharides. In addition, mannobiose and mannose were commonly detected as the hydrolyzed products of LBG, konjac and ivory nut.

• Microbiology and Biotechnology Letters
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• v.48 no.2
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• pp.158-166
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• 2020

A gene coding for the xylanase was cloned from Paenibacillus woosongensis, followed by determination of its complete nucleotide sequence. This xylanase gene, designated as xyn10A, consists of 1,446 nucleotides encoding a polypeptide of 481 amino acid residues. Based on the deduced amino acid sequence, Xyn10A was identified to be a modular enzyme composed of a catalytic domain highly homologous to the glycosyl hydrolase family 10 xylanase and a putative carbohydrate-binding module (CBM) in the C-terminus. By using DEAE-sepharose and phenyl-sepharose column chromatography, Xyn10A was purified from the cellfree extract of recombinant Escherichia coli carrying a P. woosongensis xyn10A gene. The N-terminal amino acid sequence of the purified Xyn10A was identified to exactly match the sequence immediately following the signal peptide predicted by the Signal5.0 server. The purified Xyn10A was a truncated protein of 33 kDa, suggesting the deletion of CBM in the C-terminus by intracellular hydrolysis. The purified enzyme had an optimum pH and temperature of 6.0 and 55-60℃, respectively, with the kinetic parameters V_max and K_m of 298.8 U/mg and 2.47 mg/ml, respectively, for oat spelt xylan. The enzyme was more active on arabinoxylan than on oat spelt xylan and birchood xylan with low activity for p-nitrophenyl-β-xylopyranoside. Xylanase activity was significantly inhibited by 5 mM Cu²⁺, Mn²⁺, and SDS, and was noticeably enhanced by K⁺, Ni²⁺, and Ca²⁺. The enzyme could hydrolyze xylooligosaccharides larger than xylobiose. The predominant products resulting from xylooligosaccharide hydrolysis were xylobiose and xylose.