• Title/Summary/Keyword: MsrB3

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Expression, subcellular localization, and antioxidant role of mammalian methionine sulfoxide reductases in Saccharomyces cerevisiae

  • Kwak, Geun-Hee;Kim, Jae-Ryong;Kim, Hwa-Young
    • BMB Reports
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    • v.42 no.2
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    • pp.113-118
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    • 2009
  • Despite the growing body of evidence suggesting a role for MsrA in antioxidant defense, little is currently known regarding the function of MsrB in cellular protection against oxidative stress. In this study, we overexpressed the mammalian MsrB and MsrA genes in Saccharomyces cerevisiae and assessed their subcellular localization and antioxidant functions. We found that the mitochondrial MsrB3 protein (MsrB3B) was localized to the cytosol, but not to the mitochondria, of the yeast cells. The mitochondrial MsrB2 protein was detected in the mitochondria and, to a lesser extent, the cytosol of the yeast cells. In this study, we report the first evidence that MsrB3 overexpression in yeast cells protected them against $H_2O_2$-mediated cell death. Additionally, MsrB2 overexpression also provided yeast cells with resistance to oxidative stress, as did MsrA overexpression. Our results show that mammalian MsrB and MsrA proteins perform crucial functions in protection against oxidative stress in lower eukaryotic yeast cells.

Inhibition of methionine sulfoxide reduction by dimethyl sulfoxide

  • Kwak, Geun-Hee;Choi, Seung-Hee;Kim, Jae-Ryong;Kim, Hwa-Young
    • BMB Reports
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    • v.42 no.9
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    • pp.580-585
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    • 2009
  • Dimethyl sulfoxide (DMSO) is widely used in chemistry and biology as a solvent and as a cryoprotectant. It is also used as a pharmaceutical agent for the treatment of interstitial cystitis and rheumatoid arthritis. Previous reports described DMSO as being reduced by methionine-S-sulfoxide reductase (MsrA). However, little is known about the DMSO reduction capability of methionine-R-sulfoxide reductase (MsrB) or its effect on the catalysis of methionine sulfoxide reduction. We show that mammalian MsrB2 and MsrB3 were unable to reduce DMSO. This compound inhibited MsrB2 activity but did not inhibit MsrB3 activity. We further determined that DMSO functions as an inhibitor of MsrA and MsrB2 in the reduction of methionine sulfoxides via different inhibition mechanisms. DMSO competitively inhibited MsrA activity but acted as a non-competitive inhibitor of MsrB2 activity. Our study also demonstrated that DMSO inhibits in vivo methionine sulfoxide reduction in yeast and mammalian cells.

Overexpression, Purification, and Preliminary X-Ray Crystallographic Studies of Methionine Sulfoxide Reductase B from Bacillus subtilis

  • Park, Ae-Kyung;Shin, Youn-Jae;Moon, Jin-Ho;Kim, Young-Kwan;Hwang, Kwang-Yeon;Chi, Young-Min
    • Journal of Microbiology and Biotechnology
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    • v.18 no.1
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    • pp.59-62
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    • 2008
  • The peptide methionine sulfoxide reductases (Msrs) are enzymes that catalyze the reduction of methionine sulfoxide back to methionine. Because of two enantiomers of methionine sulfoxide (S and R forms), this reduction reaction is carried out by two structurally unrelated classes of enzymes, MsrA (E.C. 1.8.4.11) and MsrB (E.C. 1.8.4.12). Whereas MsrA has been well characterized structurally and functionally, little information on MsrB is available. The recombinant MsrB from Bacillus subtilis has been purified and crystallized by the hanging-drop vapor-diffusion method, and the functional and structural features of MsrB have been elucidated. The crystals belong to the trigonal space group P3, with unit-cell parameters a=b=136.096, $c=61.918{\AA}$, and diffracted to $2.5{\AA}$ resolution using a synchrotron-radiation source at Pohang Light Source. The asymmetric unit contains six subunits of MsrB with a crystal volume per protein mass $(V_M)\;of\;3.37{\AA}^3\;Da^{-1}$ and a solvent content of 63.5%.

Optimum Combination of Pickup Coil Type and Magnetically Shielded Room for Maximum SNR to Measure Biomagnetism (생체신호 측정을 위한 최대의 신호 대 잡음비를 가지는 검출코일의 형태 와 자기차폐실의 최적 조합)

  • Yu, K.K.;Lee, Y.H.;Kang, C.S.;Kim, J.M.;Park, Y.K.
    • Progress in Superconductivity
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    • v.9 no.1
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    • pp.45-49
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    • 2007
  • We have investigated the optimum combination of the environmental noise condition and type of SQUID pickup coil in order to obtain maximum signal-to-noise ratio (SNR). The measurement probe consists of 1st order gradiometer with pickup coils of 100 mm, 70 mm, and 50 mm baseline length, a 2nd order gradiometer with 50 mm baseline, and a magnetometer. The pickup coils are fabricated by winding Nb wire on a bobbin with 200 mm diameter. Noise and heart signal of a healthy male were measured by various SQUID sensors with different types of pickup coils in various magnetically shielded rooms (MSR), and compared to each other. The shielding factors were found to be 43 dB, 35 dB and 25 dB at 0.1 Hz for MSR-AS, MSR-BS, MSR-CS, respectively. White noises were $3.5\;fT/Hz^{1/2}$, $4.5\;fT/Hz^{1/2}$ and $3\;fT/Hz^{1/2}$ for the 1st order gradiometers, the 2nd order gradiometers, and magnetometer for all MSRs. SNR of the magnetometer was up to 56 dB in MSR-AS, while the 1st order axial gradiometer with 70 mm baseline length was up to 54 dB in MSR-BS. The 2nd order axial gradiometer with 50 mm baseline length of pickup coil was found to be up to 40 dB in MSR-CS.

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Identification of an antimicrobial peptide from human methionine sulfoxide reductase B3

  • Kim, Yong-Joon;Kwak, Geun-Hee;Lee, Chu-Hee;Kim, Hwa-Young
    • BMB Reports
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    • v.44 no.10
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    • pp.669-673
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    • 2011
  • Human methionine sulfoxide reductase B3A (hMsrB3A) is an endoplasmic reticulum (ER) reductase that catalyzes the stereospecific reduction of methionine-R-sulfoxide to methionine in proteins. In this work, we identified an antimicrobial peptide from hMsrB3A protein. The N-terminal ER-targeting signal peptide (amino acids 1-31) conferred an antimicrobial effect in Escherichia coli cells. Sequence and structural analyses showed that the overall positively charged ER signal peptide had an Argand Pro-rich region and a potential hydrophobic ${\alpha}$-helical segment that contains 4 cysteine residues. The potential ${\alpha}$-helical region was essential for the antimicrobial activity within E. coli cells. A synthetic peptide, comprised of 2-26 amino acids of the signal peptide, was effective at killing Gram-negative E. coli, Klebsiella pneumoniae, and Salmonella paratyphi, but had no bactericidal activity against Gram-positive Staphylococcus aureus.

Detection of Inducible Clindamycin Resistance Genes (ermA, ermB, and ermC) in Staphylococcus aureus and Staphylococcus epidermidis

  • Mazloumi, Mohammad Javad;Akbari, Reza;Yousefi, Saber
    • Microbiology and Biotechnology Letters
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    • v.49 no.3
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    • pp.449-457
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    • 2021
  • The aim of the present study was to survey the frequency of inducible and constitutive phenotypes and inducible cross-resistant genes by regulating the methylation of 23S rRNA (ermA, ermB, and ermC) and macrolide efflux-related msrA gene in Staphylococcus aureus and S. epidermidis strains. A total of 172 bacterial isolates (identified based on standard tests), were examined in this study. Antibiotic susceptibility was determined by the disk diffusion method, and all isolates were evaluated with respect to inducible and constitutive phenotypes. The presence of ermA, ermB, ermC, and msrA genes was investigated by a PCR assay. The constitutive resistance phenotypes showed a higher distribution among the isolates. R phenotype was detected more among S. epidermidis isolates (46.25%). ermB, ermC, and msrA genes were detected more in methicillin-resistant S. aureus (MRSA) and methicillin-resistant S. epidermidis (MRSE) isolates that had R and HD phenotypes (>77% strains). The ermA gene had the lowest frequency among MRSA, MRSE, MSSA, and MSSE strains (<14% isolates). Distribution of inducible resistance genes in MRSA and MRSE strains, and possibly other species, leads to increased constitutive resistance to erythromycin, clindamycin, and other similar antibiotics. Therefore, it can be challenging to treat infections caused by these resistant strains.

Resistance to Macrolide, Lincosamide and Streptogramin Antibiotics in Staphylococci Isolated in Istanbul, Turkey

  • Aktas, Zerrin;Aridogan, Aslihan;Kayacan, Cigdem Bal;Aydin, Derya
    • Journal of Microbiology
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    • v.45 no.4
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    • pp.286-290
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    • 2007
  • The purpose of this study was to investigate the prevalence and genetic mechanisms of erythromycin resistance in staphylococci. A total of 102 erythromycin resistant non-duplicate clinical isolates of staphylococci [78. coagulase negative stapylococci (CNS), 24 Staphylococcus aureus] were collected between October 2003 and August 2004 in Istanbul Faculty of Medicine in Turkey. The majority of the isolates were from blood and urine specimens. Antimicrobial susceptibilities were determined by the agar dilution procedure and the resistance phenotypes by the double disk induction test. A multiplex PCR was performed, using primers specific for erm(A), erm(B), erm(C), and msrA genes.. Among the 78 CNS isolates, 57.8% expressed the $MLS_{B}-constitutive$, 20.6% the $MLS_{B}-inducible$, and 21.6% the $MS_B$ phenotypes. By PCR, 78.2% of these isolates harbored the erm(C) gene, 8.9% erm(A), 6.4% erm(B), and 11.5% msrA genes. In S. aureus, the constitutive $MLS_B$ (58.3 %) was more common than the inducible phenotype (20.8%). erm(A) was detected in 50% and erm(C) in 62.5% of the isolates, while 37.5% contained both erm(A) and erm(C). erm(C)-associated macrolide resistance was the most prevalent in CNS, while ermC) and erm(A, C) was the most prevalent in S. aureus.

On the Development of Parthenogenetic Oocytes by Cytochalasin B and Production of Cloned Mice by SCNT

  • Sim, Bo-Woong;Min, Kwan-Sik
    • Journal of Embryo Transfer
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    • v.29 no.2
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    • pp.111-117
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    • 2014
  • This study was conducted to optimize the efficiency of cloning and to produce cloned mice. The majority of cloned mammals derived by nuclear transfer (NT) die during gestation and have enlarged and dysfunctional placentas. In this study, the optimized conditions were established to produce clone mice. The parthenogenetic oocytes were activated after 6 h regardless of cytochalasin B (CB) concentration. CB treatment ($2{\mu}g/ml$) was found second polar body. Lower concentration of CB was decreased the activation rate, but the second polar body was the best highly increased during 6 h incubation. The small fragments were exhibited in the $5{\mu}g/ml$ treatment of CB, but it was not found in lower concentration groups (> $2.5{\mu}g/ml$). To examine effects of $SrCl_2$ on the adult cumulus cells, somatic cell NT oocytes were exposed during 0.5, 1 and 6 hrs. The second polar body was significantly greater in 0.5 h exposure group (6.6%) than 1, 6 hrs. Developmental rate from 2-cell to 4-cell was the lowest in 7.5 mM Strontium chloride ($SrCl_2$) groups (84.1% and 64.3%) than 5, 10 m $MSrCl_2$. The implantation rate was not significantly difference among 5, 7.5 and 10 m $MSrCl_2$ group. Three live fetuses were produced by SCNT. SCNT placentas were remarkably heavier than IVF group (8 fetuses) (0.34, 0.34, 0.33 vs 0.14 g) compared with the placenta weight of IVF and SCNT clones.

Synthesis of TiN/TiB2/Ti-silicides Nanocomposite Powders by Mechanochemical Reaction and its Reaction Mechanism (기계화학반응에 의한 TiN/TiB2/Ti-silicides 나노복합분말의 합성과 반응기구)

  • Cho Young-Whan;Kim Ji-Woo;Shim Jae-Hyeok;Ahn Jae-Pyoung;Oh Kyu-Hwan
    • Journal of Powder Materials
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    • v.12 no.4 s.51
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    • pp.273-278
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    • 2005
  • Nanostructured TiN/$TiB_2$/$TiSi_2$ and TiN/$TiB_2$/$Ti_5Si_2$ composite powders have been prepared by mechanochemical reaction from mixtures of Ti, BN, and $Si_3N_4$ powders. The raw materials have reacted to form a uniform mixture of TiN, $TiB_2$ and $TiSi_2$ or $Ti_5Si_3$ depending on the amount of $Si_3N_4$ used in the starting mixtures, and the reaction proceeded through so-called mechanically activated self-sustaining reaction (MSR). Fine TiN and $TiB_2$ crystallites less than a few tens of nanometer were homogeneously dispersed in the amorphous $TiSi_2$ or $Ti_5Si_3$ matrix after milling for 12 hours. These amorphous matrices became crystalline phases after annealing at high temperatures as expected, but the original microstructure did not change significantly.

Antibiotic Resistance and Safety Assessment of Enterococcus faecium CKDB003 for Using as Probiotics (프로바이오틱스 Enterococcus faecium CKDB003의 항생제 내성 및 안전성 평가)

  • Kim, Han Jun;Kang, Soon Ah
    • The Korean Journal of Food And Nutrition
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    • v.33 no.3
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    • pp.223-236
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    • 2020
  • In this study, a safety evaluation was conducted to confirm if the Enterococcus faecium CKDB003 strain obtained by selection from a mixed fermentation of fruit and milk is suitable for use as a probiotic. The MIC value for the 10 antibiotics specified in the EFSA guidance was below the acceptable cut-off value. The antibiotic resistance genes aac(6')-li, eatAv, and msr(C) exist by whole genome sequencing, but are in the chromosome and not in the plasmid, thus confirming that there is no possibility of transmission to other microorganisms. It was confirmed that cytolysin (cylA, cylB, cylI, cylL-l, cylL-s, cylM, cylR1, cylR2), aggregation substance (asa1, asp1), collagen adhesion (ace), enterococcal surface protein (esp), endocarditis antigen (efaA), hyaluronidase (hyl) and gelatinase (gelE) were not present in the genome by examining the genes of factors related to virulence. Also, the biochemical analysis showed no toxic enzyme activities, and no virulence genes were detected by the PCR method. Thus, the E. faecium CKDB003 strain can be safely used as a health functional food probiotic, based on the results of the safety assessment.