• Title/Summary/Keyword: HRC

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Reduction-Sensitive and Cysteine Residue-Mediated Streptococcus pneumoniae HrcA Oligomerization In Vitro

  • Kwon, Hyog-Young;Kim, Eun-Hye;Tran, Thao Dang Hien;Pyo, Suhk-Neung;Rhee, Dong-Kwon
    • Molecules and Cells
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    • v.27 no.2
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    • pp.149-157
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    • 2009
  • In both gram-positive and several gram-negative bacteria, the transcription of dnaK and groE operons is negatively regulated by HrcA; however, the mechanism modulating HrcA protein activity upon thermal stress remains elusive. Here, we demonstrate that HrcA is modulated via reduction and oligomerization in vitro. Native-PAGE analysis was used to reveal the oligomeric structure of HrcA. The oligomeric HrcA structure became monomeric following treatment with the reducing agent dithothreitol, and this process was reversed by treatment with hydrogen peroxide. Moreover, the mutant HrcA C118S exhibited reduced binding to CIRCE elements and became less oligomerized, suggesting that cysteine residue 118 is important for CIRCE element binding as well as oligomerization. Conversely, HrcA mutant C280S exhibited increased oligomerization. An HrcA double mutant (C118S, C280S) was monomeric and exhibited a level of oligomerization and CIRCE binding similar to wild type HrcA, suggesting that cysteine residues 118 and 280 may function as checks to one another during oligomer formation. Biochemical fractionation of E. coli cells overexpressing HrcA revealed the presence of HrcA in the membrane fraction. Together, these results suggest that the two HrcA cysteine residues at positions 118 and 280 function as reduction sensors in the membrane and mediate oligomerization upon stress.

Antioxidant Property and Inhibitory Effects of an Water Extract of Hwang-Ryun-Chung-Sim-Um on the Acetylcholinesterase (황련청심음(黃連淸心飮)의 항산화 및 AChE 억제 효과에 관한 연구)

  • Yoo, Jong-Ho;Lee, Sang-Taek;Han, Yun-Seung;Kim, Geun-Woo;Koo, Byung-Soo;Kim, Hun-Il
    • Journal of Oriental Neuropsychiatry
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    • v.17 no.1
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    • pp.1-16
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    • 2006
  • Objective: An water extract of the Hwang-Ryun-Chung-Sim-Um (HRC) was assessed to determine the mechanisms of its antioxidant activity. In addition, the HRC was examined in vitro for the inhibitory effect on the acetylcholinesterse (AChE). Methods: The HRC exhibited a concentration-treatment; scavenging ${\alpha},{\alpha}-diphenyl-{\beta}-picrylhydrazyl$ (DPPH) radical, linoleic acid oxidation in a thiocyanate assay system, hydroxyl radical-induced DNA nicking. We investigated mRNA levels such as catalase activity, superoxide-dismutase and glutathione peroxidase. The water extract of HRC showed inhibitory effect on AChE activity. Result: The HRC extract showed dose-dependent free radical scavenging activity, including DPPH radicals and hydroxyl radicals, using different system. The HRC was also found to be effective in protecting plasmid DNA against the strand breakage induced by Hydroxyl radicals in Fenton's reaction mixture. Futhermore, catalase mRNA expression levels increased, but SOD1 and MnSOD was not expressed. HRC in a various concentration-dependent decreased AChE mRNA levels and inhibitory effect showed AChE. Conclusion: According to the above results, it is supposed that HRC is applicable to the Dementia-type of Alzheimer clinically.

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Characterization of Ca2+-Dependent Protein-Protein Interactions within the Ca2+ Release Units of Cardiac Sarcoplasmic Reticulum

  • Rani, Shilpa;Park, Chang Sik;Sreenivasaiah, Pradeep Kumar;Kim, Do Han
    • Molecules and Cells
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    • v.39 no.2
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    • pp.149-155
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    • 2016
  • In the heart, excitation-contraction (E-C) coupling is mediated by $Ca^{2+}$ release from sarcoplasmic reticulum (SR) through the interactions of proteins forming the $Ca^{2+}$ release unit (CRU). Among them, calsequestrin (CSQ) and histidine-rich $Ca^{2+}$ binding protein (HRC) are known to bind the charged luminal region of triadin (TRN) and thus directly or indirectly regulate ryanodine receptor 2 (RyR2) activity. However, the mechanisms of CSQ and HRC mediated regulation of RyR2 activity through TRN have remained unclear. We first examined the minimal KEKE motif of TRN involved in the interactions with CSQ2, HRC and RyR2 using TRN deletion mutants and in vitro binding assays. The results showed that CSQ2, HRC and RyR2 share the same KEKE motif region on the distal part of TRN (aa 202-231). Second, in vitro binding assays were conducted to examine the $Ca^{2+}$ dependence of protein-protein interactions (PPI). The results showed that TRN-HRC interaction had a bell-shaped $Ca^{2+}$ dependence, which peaked at pCa4, whereas TRN-CSQ2 or TRN-RyR2 interaction did not show such $Ca^{2+}$ dependence pattern. Third, competitive binding was conducted to examine whether CSQ2, HRC, or RyR2 affects the TRN-HRC or TRN-CSQ2 binding at pCa4. Among them, only CSQ2 or RyR2 competitively inhibited TRN-HRC binding, suggesting that HRC can confer functional refractoriness to CRU, which could be beneficial for reloading of $Ca^{2+}$ into SR at intermediate $Ca^{2+}$ concentrations.

Regulation of Contraction and $Ca^{2+}$ Transient by Histidine-rich $Ca^{2+}$-binding Protein in Ventricular Myocytes (히스티딘-리치 $Ca^{2+}$ 결합 단백질에 의한 심실근세포 수축 및 $Ca^{2+}$ Transient의 조절)

  • Son, Min-Jeong;Kim, Joon-Chul;Kim, Seong-Woo;Ahn, Jong-Real;Woo, Sun-Hee
    • YAKHAK HOEJI
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    • v.56 no.6
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    • pp.377-381
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    • 2012
  • The histidine-rich $Ca^{2+}$ binding protein (HRC) is a $Ca^{2+}$ binding protein in the sarcoplasmic reticulum (SR). In this study, we examined whether the HRC is involved in the regulation of cardiac contraction and $Ca^{2+}$ signaling using HRC knock-out (KO) mouse ventricular myocytes. In field-stimulated single mouse ventricular myocytes, cell shortenings and $Ca^{2+}$ transients were measured using a video edge detection and a confocal $Ca^{2+}$ imaging, respectively. Compared with the wide-type (WT) myocytes, the magnitudes of cell shortenings were significantly larger in HRC KO cells (P<0.01, WT vs. KO). The rate of contraction and relaxation was significantly accelerated in HRC KO myocytes (P<0.05 and P<0.01, respectively, WT vs. KO). The magnitudes of $Ca^{2+}$ transients were increased by HRC KO (P<0.01, WT vs. KO). In addition, the decay of the $Ca^{2+}$ transient was faster in HRC KO cells than in wild-type cells P<0.01, WT vs. KO). These results suggest that HRC may suppress SR $Ca^{2+}$ releases and decay of $Ca^{2+}$ transients during action potentials, thereby attenuating ventricular contraction and relaxation.

Differential Regulation of the Genes of the Streptococcus pneumoniae dnaK Operon by Ca++

  • Kim, Seung-Whan;Bae, Yong-Goo;Pyo, Suhk-Neung;Rhee, Dong-Kwon
    • Molecules and Cells
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    • v.23 no.2
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    • pp.239-245
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    • 2007
  • DnaK is a major antigen in Streptococcus pneumoniae, and is induced by a minor shift in temperature (30 to $37^{\circ}C$) but not by ethanol shock. Although HrcA in the presence of $Ca^{{+}{+}}$ represses the expression of both groEL and hrcA, the control of transcription of the dnaK operon is not completely understood. In this study, the dnaK operon of S. pneumoniae (5' hrcA-grpE-dnaK-dnaJ) was cloned and analyzed. It contains large intergenic regions in grpE/dnaK and dnaK/dnaJ. Pulse labeling with [$^{35}S$]-methionine and immunoblot analyses revealed the presence of higher levels of DnaK than of HrcA even in the presence of $Ca^{{+}{+}}$ after heat shock suggesting that $Ca^{{+}{+}}$ differentially regulates the heat shock responses of hrcA and dnaK. By blocking de novo mRNA synthesis with rifampin it was shown that neither the hrcA nor the groEL transcripts were stabilized by heat shock even though dnaK transcripts were stabilized. We conclude that S. pneumoniae uses fine regulation of the transcription of the individual genes of the tetracistronic dnaK operon to cope with the various stresses experienced during infections.

Effects of Rosa multiflora and Rosa multiflora Complex on Lipid Content in Rats Fed a High-Fat·High-Cholesterol Diet (찔레꽃 및 찔레꽃 복합물 추출물이 고지방·고콜레스테롤 식이 흰쥐의 지질 함량 개선에 미치는 영향)

  • Choi, Jeong-Hwa;Baek, Ji-Yun;Choi, Hee-Jin
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.44 no.6
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    • pp.791-799
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    • 2015
  • This study was conducted to investigate the effects of water extract from Rosa multiflora and Rosa multiflora complex on lipid metabolism in rats fed a high cholesterol diet. Experimental rats were divided into four groups, which were composed of normal diet group (N group), high cholesterol diet group (HF group), high cholesterol and 5% Rosa multiflora diet group (HR group), and high cholesterol and Rosa multiflora complex diet group (HRC group). Rosa multiflora complex is composed of Duchesnea chrysantha, Salvia plebeia R. BR., and Sasa borealis. The serum triglyceride (TG) content of the HF group was significantly higher than that of the N group, whereas that of the HRC group was significantly lower. Serum high-density lipoprotein-cholesterol contents of the HR and HRC groups were significantly higher than that of the HF group. The serum total cholesterol and low-density lipoprotein- cholesterol contents of the HF group were significantly higher than those of the N group, and especially the atherogenic index of the HRC group was significantly reduced compared to the HF group. The liver TG and total cholesterol contents of the HF group were significantly increased compared to the normal diet group, while those of the HR and HRC groups were significantly decreased compared to the HF group. Fecal total lipid and total cholesterol contents of the HR and HRC groups were significantly increased compared to the HF group. Uridine 5'-diphospho (UDP)-glucuronyl transferase activity of the HR and HRC groups were increased compared to the HF group. Total bile acid contents of the HR and HRC groups were increased compared to the HF group, and that of the HRC group was significantly increased compared to the HF groups. These results suggest that Rosa multiflora supplementation has powerful health benefits due to UDP-glucuronyl transferase activity, bile acid, and lipid metabolism.

Characteristics of Carbozymethylated Substrates from Delignified Autohydrolyzed Substrates (탈리그닌한 자기가수분해 시료로부터 준비한 카복시메틸화 시료의 특성)

  • Cho, Nam-Seok
    • Journal of the Korean Wood Science and Technology
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    • v.32 no.1
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    • pp.28-34
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    • 2004
  • This study was performed to evaluate the characteristics of the carboxymethylated substrate from high reactive autohydrolyzed cellulose (HRC) and those of commercial α-cellulose (CAC) and refiner mechanical pulp (RMP). Saccharification rates of HRC substrate were achieved over 70% with 12 hr hydrolysis, about 90% with 24 hr, and 99.5% with 72 hr. CMCase and avicelase activities of cellulase onozuka were 4.09 ㎛ G/mg·min and 14.0 ㎛ G/mg·min, respectively. There were no any significant changes in cellulase activities with this substrate. The saccharification rates of CAC and RMP were very low, 57% and 38% with 72 hr, respectively. Those lignin-zero autohydrolyzed substrates, HRC and CAC, were highly carboxymethylated at the high alkali concentration, near 30%, for 3 hr. reaction, and resulted in 1.13-1.15 of D.S., besides 0.85 of D.S. from RMP. Water solubilities of carboxymethylated substrates were increased with an increase of D.S., 98-98.5% from HRC and CAC and 31.5% from RMP. RMP which has low specific surface area showed lower water retention values, compared to high values of 435 and 321% from CAC and HRC, respectively. There were no direct relationship between surface area and swelling ratio of the substrates.

An Experimental Study on Structural Performance of HRC Composite Beam according to types of Connection Plate with Stud Bolts (HRC 복합보의 연결플레이트 보강법에 따른 구조성능실험)

  • Lee, Soo-Kueon;Yang, Jae-Guen;Song, Chang-Seok;Jang, Eun-Young;Moon, Jun-Chul
    • Journal of Korean Society of Steel Construction
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    • v.23 no.3
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    • pp.377-384
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    • 2011
  • Recently, for the purpose of reducing work terms and improving performance for construction work, various methods in structure field were developed. This included the HRC system which is applicable to a typical structure (e.g., parking and office building). The HRC system introduced the Gerber Joint to raise structural efficiency and used connection plate to bolt HRC composite beam to H beam in the construction field. In this research, the experimental tests for six specimens, which were in the same field conditions, were conducted with several parameters such as the length and height of the connection plate and the number of stub bolts. The test result was compared with those made by current design codes for the deflection and strains of the main bar. Within the given load, the integration of concrete in beam and connection plate, welded with stud bolts, was verified.