• YAKHAK HOEJI
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• v.42 no.1
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• pp.53-58
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• 1998

A novel type of sulfotransferase, arylsulfate sulfotransferase (EC 2.8.2.22) purified from Haemophilus K-12, an intestinal bacterium of a mouse, was immobilized onto AH-S epharose 4B, CH-Sepharose 4B and DEAE-celluose. The enzyme was stabilized for storage more markedly by covalent immobilization onto AH-Spharose 4B or CH-Sepharose 4B and by adsorptive immobilization onto DEAE-cellulose than the free enzyme. The optimal pH and acceptor substrate specificity of immobuized enzyme were similar to those of the free enzyme.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1994.04a
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• pp.257-257
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• 1994

생쥐의 장내세균으로 부터 황산전이 반응을 촉매하는 효소인 sulfotransferase를 생산하는 균주를 분리하였으며 동정결과 Haemophilus속 균주로 확인되어 Haemophilus K-12라 명명하였다. 균의 성장과 효소활성과의 관계를 보면 균은 10시간에서 완전히 성장하였으며 효소활성도 이와 비례하였다. Haemophilus K-12의 배지조성에 따른 sulfotransferase의 생산성을 Brain Heart Infusion 배지에서의 생산성과 비교해보면 탄소원으로는 sucrose가 0.2%농도에서 584%로 가장 좋았으며 질소원으로는 yeast extract가 266%로 가장 좋았다. 공여체로 PNS를 최종농도 1mM로 하여 배지에 첨가하였을때 212%로 가장 높은 효소증가를 보였다. 2가금속이온에 의한 효소증가는 현저하지 않았으며 $Mn^{2+}$이 107%로 가장 높았고$Zn^{2+}$와 EDTA에 의해서는 저해를 받았다. 이상의 결과를 종합하여 균배양을 위한 이상적인 배지조성을 sucrose 0.2%, yeast extract 1%, $Na_2$HPO$_4$ 0.25%, NaCl 0.5%로 결정하였다. 결정된 최적배지에 균을 10L 배양하여 초음파 처리, 원심분리한 것을 70 % ammonium sulfate fractionation, DEAE-cellulose column chromatography를 2회, Hydroxyapatite column chromatography, chromatdfocusing column chromatography, Silica PAE column chromatography, Sephacryl S-300 superfine column chromatography를 행한결과 specific activity가 6.76 umoie/min/mg protein인 효소액을 얻었으며 homogeneous enzyme였다. 이렇게 해서 얻은 효소를 이용하여 수용체 기질 특이성을 측정한 결과 1-naphthol이 phenol을 100%로 하였을 때 233%로 가장 좋았으며 Eubacterium A-44 sulfotransferase의 좋은 기질이었던 p-acetaminophenol, tyramine, 9-phenanthrol은 좋은 기질이 되지 못했다. 이상의 결과로 미루어 보아Haemophilus K-12 sulfotransferase는 지금까지 보고된 bacterial sulfotrasferase와는 다른 효소로 사려되며 효소반응기전의 규명이 이루어지면 산업적 응용이 가능할것으로 사료된다.

• YAKHAK HOEJI
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• v.36 no.5
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• pp.455-459
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• 1992

Haemophilus K-12 producing novel sulfotransferase was isolated from mouce intestinal flora. The enzyme catalyzed the transfer of sulfate group from p-nitrophenylsulfate to phenolic compounds. The optimum medium condition for the sulfotransferase production was 0.2% sucrose, 1% yeast extract, $Na_{2}HPO_4$ and 0.5% NaCl. The enzyme production was induced by donor substrate, but was not by accepters. When p-nitrophenylsulfate was used as a donor substrate, 1-naphthol was best substrate, followed by phenol, p-acetaminophenol and tyramine.

• Journal of Microbiology and Biotechnology
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• v.19 no.12
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• pp.1582-1589
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• 2009

Bacterial UDP-N-acetylglucosamine enolpyruvyl transferase (MurA) catalyzes the transfer of enolpyruvate from phosphoenolpyruvate (PEP) to uridine diphospho-N-acetylglucosamine (UNAG), which is the first step of bacterial cell wall synthesis. We identified thimerosal, thiram, and ebselen as effective inhibitors of Haemophilus influenzae MurA by screening a chemical library that consisted of a wide range of bioactive compounds. When MurA was preincubated with these inhibitors, their 50% inhibitory concentrations ($IC_{50}s$) were found to range from 0.1 to $0.7\;{\mu}M$. In particular, thimerosal suppressed the growth of several different Gram-negative bacteria such as Escherichia coli, Pseudomonas aeruginosa, and Salmonella typhimurium at a concentration range of $1-2\;{\mu}g/ml$. These inhibitors covalently modified the cysteine residue near the active site of MurA. This modification changed the open conformation of MurA to a more closed configuration, which may have prevented the necessary conformational change from occurring during the enzyme reaction.

• Molecules and Cells
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• v.19 no.3
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• pp.398-401
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• 2005

The bacterial enzyme UDP-N-acetylglucosamine enolpyruvyl transferase catalyzes the first committed step of peptidoglycan biosynthesis, i.e., transfer of enolpyruvate from phosphoenolpyruvate to UDP-N-acetyl-glucosamine. We have overexpressed the enzyme from Haemophilus influenzae in Escherichia coli and crystallized it in the apo-form, as well as in a complex with UDP-N-acetylglucosamine and fosfomycin using ammonium sulfate as the precipitant. X-ray diffraction data from a crystal of the apo-form were collected to $2.8{\AA}$ resolution at 293 K. The crystal quality was improved by co-crystallization with UDP-N-acetylglucosamine and fosfomycin. X-ray data to $2.2{\AA}$ have been collected at 100 K from a flash-frozen crystal of the complex. The complex crystals belong to the orthorhombic space group I222 (or $I2_12_12_1$) with unit-cell parameters of a = 63.7, b = 124.5, and $c=126.3{\AA}$. Assuming a monomer of the recombinant enzyme in the crystallographic asymmetric unit, the calculated Matthews parameter ($V_M$) is $2.71{\AA}^3Da^{-1}$ and solvent content is 54.6%.

• Bulletin of the Korean Chemical Society
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• v.31 no.12
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• pp.3782-3784
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• 2010

• Proceedings of the Korean Society of Applied Pharmacology
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• 1993.04a
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• pp.142-142
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• 1993

생쥐에서 sulfotransferase 생산 군주를 검색한 결과 호기성균보다 혐기성균이 많은 것으로 나타났다. 이러한 것은 전번 보고와 일치한다. 황산전이효소 생산 균주 검색용 배지에서 형광을 나타내는 호기성 균주를 분리하여 K-12라 명하였다. 동정한 결과 K-12균주는 그람 음성으로 운동성이 없는 통성혐기성 균주인 Haemophilus sp.로 사료된다. 이 균주의 황산전이효소 생산성은 공여체 기질만을 배지에 첨가했을때 가장 높은 생산성을 보였다. 탄소원으로는 sucrose와 lactose를 사용했을때 가장 높은 효소유도를 보였으며, 질소원으로는 yeast extract, peptone이 우수하였다. 배양최적 pH는 7부근이었으며 온도는 37$^{\circ}C$가 가장 좋았다. 금속이온으로는 $Mg^{2＋}$이 우수했다. 최적 배지 10 L에 K-12 균주를 배양하여, 집균 초음파 처리, 원심분리한 상등액을 70% ammonium sulfate fractionation, DEAE-cellulose column chromatography 2회, Sephcryl S-300 superfine column chromatography를 행한 결과 specific activity가 0.267 $\mu$mole/min/mg protein인 효소를 부분 정제하였다.

• Clinical and Experimental Pediatrics
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• v.50 no.5
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• pp.449-456
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• 2007

Purpose : Antibody persistence after primary series of Haemophilus influenzae type b (Hib) vaccine and responses to a boosters are little known in Korean children. We performed this study to evaluate the antibody titer in relation with a booster immunization of Hib vaccine in Korean children. Methods : One hundred forty-four children aged 12-23 months old were enrolled in three university hospitals. The immunogenicity of a boosters with Hib vaccine was assessed in children previously primed with Hib vaccine. Antibody persistence was also assessed in children who had received 3 doses of Hib vaccine without a booster. Anti-polyribosylribitol phosphate (PRP) IgG antibody levels and bactericidal titers were determined by enzyme immunoassay and bactericidal assay at the Center for Vaccine Evaluation and Study, Medical Research Institute, Ewha Womans University. Results : Prior to a booster in the second year of life, geometric mean antibody concentrations were $2.39{\mu}g/mL$ and the percent of subjects who had a anti-PRP antibody level ${\geq}1{\mu}g/mL$ was 68.6%. After boosting, antibody concentration was $19.09{\mu}g/mL$ and the percent of subjects who had a anti-PRP antibody level ${\geq}1{\mu}g/mL$ was 96.5%, which reflects previous immune priming. In subjects who had finished primary immunization only, the bactericidal titer was 3,946 and in subjects who had a booster, it was 11,205. Anti-PRP antibody level was correlated with serum bactericidal titer. Conclusion : Many children aged 12-23 month old still had protective antibodies after recommended primary immunization only. A booster dose seemed to induce good anamnestic antibody responses in Korean children.

• Microbiology and Biotechnology Letters
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• v.33 no.2
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• pp.142-147
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• 2005

The objective of this study was to develop a new potent antibacterial compound against bovine pneumoniae bacteria from medicinal plants or herbs. Among 65 kinds of medicinal plants and herbs, ethanol extracts of Citrus unishiu showed the highest solid yield of $54\%$. However, ethanal extracts from Agastache rugosa had the highest antibacterial activities against bovine pneumoniae bacteria, Mannheimia haemolytica A and Haemophilus somnus (size of clear zone: 16.0 mm and 10.0 mm, respectively). The antibacterial compound was also maximally extracted when the powder of A. rugosa was treated with $70\%$ ethanol at $45^{\circ}C$ for 12 hours.

• Pediatric Infection and Vaccine
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• v.13 no.2
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• pp.124-129
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• 2006

Purpose : We investigated the causative organisms of bacterial meningitis by age distribution from 1996 to 2005. Methods : Retrospective data were obtained from the medical records with diagnosis of bacterial meningitis or neonatal meningitis from 1996 through 2005. A case was defined by isolation of organism or detection of its antigen by latex agglutination from cerebrospinal fluid. Results : A total of 46 cases(27 neonates and 19 children) were identified. 15 of 27 episodes(55.6%) of neonatal meningitis had a concomitant-positive blood culture. Group B streptococci were the most common bacterial causes of neonatal meningitis(44.4%). Nine of 12 episodes(75.0%) were late-onset infections in neonatal meningitis caused by group B streptococci. 16 of 19 children(84.2%) with bacterial meningitis beyond the neonatal period were younger than 5 years of age(median age, 23 months). Of 19 cases, 8 infections were with Streptococcus pneumoniae, 8 were with Haemophilus influenzae and 3 were with Neisseria meningitidis. Since 2001 there was no case of meningococcal meningitis in this study. Conclusion : In neonates group B streptococci are the most common causative organisms of bacterial meningitis, especially late-onset infections. In infants and young children, the predominant causes of bacterial meningitis are H. influenzae and S. pneumoniae; meningitis caused by the former are likely to decrease after the introduction of the conjugate vaccine for H. influenzae type b.