• 통합자연과학논문집
• /
• 제6권1호
• /
• pp.57-63
• /
• 2013

Histone deacetylase (HDACs) is an enzyme family that deacetylates histones and non-histones protein. Availability of crystal structure of HDAC8 has been a boosting factor to generate target based inhibitors. Hydroxamic class is the most studied one to generate potent inhibitors. HDAC class I and class II enzymes are emerging as a therapeutic target for cancer, diabetes, inflammation and other diseases. DNA methylation and histone modification are epigenetic mechanism, is important for the regulation of cellular functions. HDACs enzymes play essential role in gene transcription to regulate cell proliferation, migration and death. The aim of this article is to provide a comprehensive overview about structure and function of HDACs enzymes, histone deacetylase inhibitors (HDACi) and HDACs enzymes as a therapeutic target for cancer, inflammation and diabetes.

• The Korean Journal of Physiology and Pharmacology
• /
• 제15권6호
• /
• pp.363-369
• /
• 2011

The present study elucidated the effect of the selective inducible nitric oxide synthase (iNOS) inhibitor $N^6$-(1-iminoethyl)-L-lysine (L-NIL) on monosodium urate (MSU) crystal-induced inflammation and edema in mice feet. L-NIL (5 or 10 mg/kg/day) was administered intraperitoneally 4 h before injection of MSU (4 mg) into the soles of mice hindlimb feet. Twenty-four hours after MSU injection, foot thickness was increased by 160% and L-NIL pretreatment reduced food pad swelling in a dose dependent manner. Pretreatment of 10 mg/kg/day L-NIL significantly suppressed the foot pad swelling by MSU. Plasma level of nitric oxide (NO) metabolites and gene expression and protein level of iNOS in feet were increased by MSU, which was suppressed by L-NIL pretreatment. Similar pattern of change was observed in nitrotyrosine level. MSU increased the gene expression of tumor necrosis factor (TNF)-${\alpha}$ and interleukin (IL)-$1{\beta}$ and L-NIL pretreatment suppressed MSU-induced cytokines expression. The mRNA levels of superoxide dismutase and glutathione peroxidase1 were increased by MSU and L-NIL pretreatment normalized the gene expression. Phosphorylation of extracellular signal-regulated kinase 1/2 and p38 was increased by MSU, which was suppressed by L-NIL pretreatment. The mRNA levels of iNOS, TNF-${\alpha}$, and IL-$1{\beta}$ were increased by MSU in human dermal fibroblasts, C2C12 myoblasts, and human fetal osteoblasts in vitro, which was attenuated by L-NIL in a dose dependent manner. This study shows that L-NIL inhibits MSU-induced inflammation and edema in mice feet suggesting that iNOS might be involved in MSU-induced inflammation.

• Asian Pacific Journal of Cancer Prevention
• /
• 제16권14호
• /
• pp.5635-5641
• /
• 2015

Background: Cirrhosis is regarded as a possible end stage of many liver diseases, including viral infection. It occurs when healthy liver tissue becomes damaged and is replaced by scar tissue and finally may lead to hepatocellular carcinoma. Interferons (IFNs)are two general categories, type I and II. Type I includes one beta interferon and over 20 different alpha interferons. Alpha interferons are very similar in how they work, interacting with other proteins on cells like receptors. The main objective of this study was to compare Mx gene productivity post different cell line treatment with imported and Egyptian biosimilar locally produced IFNs, as well as the efficacy of those tested IFNs. Also, an assessment was made of sensitivity of different cell lines as alternatives to that recommended for evaluation of antiviral activity. Materials and Methods: Different cell lines (Vero, MDBK and Wish) were employed to evaluate cytotoxicity using the MTT assay. Antiviral activity was evaluated compared with standard IFN against VSV, Indiana strain -156, on tested rh-IFNs (imported; innovated and Egyptian biosimilar locally produced IFNs) in the pre-treated cell lines previously mentioned. The virus was propagated in the Wish cell line as recommended. Finally we estimated up-regulation of the Mx gene as a biomarker. Results: Data recorded revealed that test IFNs were safe in test cell lines. Viability was around 100%. Locally tested interferon did not realize the international potency limits, while the imported one was accepted compared with the standard IFN. These results were the same either using infectivity titer reduction assay or crystal violet staining of residual non- infected cells. Mx protein production was cell type related and confirmed by the detected Mx gene expressed in imported and locally produced IFN pre-treated cell lines. The expression of the gene was arranged in the order of Vero> wish > MDBK for the imported IFN, while for the Egyptian biosimillar locally produced one it was MDBK> Vero> wish. With regard to the antiviral activity there was a significant difference of imported IFN potency compared with the locally produced IFN (P<0.05), the IFN potential (antiviral activity) was not cell line related and showed non-significant difference for each separate product. Conclusions: Vero cells can be used as an alternative cell line for evaluation of IFN potency in case of unavailable USP recommended cell lines. Alternative potency evaluation assay could be used and proved significant difference in IFN potency in case of local and imported agents. Evaluation of antiviral activity could be used in parallel to viral infectivity reduction assay for better accuracy. Mx gene can be used as a marker for IFN potential.

• KSBB Journal
• /
• 제19권1호
• /
• pp.42-49
• /
• 2004

Racemic epoxide에 대한 입체선택적 가수분해능을 가지고 있는 곰팡이, Aspergillus niger LK로부터 epoxide hydrolase (EH, EC 3.3.2.3) 유전자의 진화적 유연관계 분석을 행하였다. A. niger LK의 EH 염기서열로부터 유추한 EH 단백질 아미노산 서열은 여러 박테리아의 EH들 및 포유동물의 microsomal EH들과 유의적인 유사성을 가지고 있었으며 a/$\beta$ hydrolase fold family에 속하였다. A. niger LK의 EH 단백질의 입체구조예측은 Protein Data Bank에 수록된 lqo7의 3D 결정구조와 90.6% identity를 가지는 것으로 나타났으며 다른 EH들의 아미노산 서열비교를 행한 결과 Asp$^{192}$ , Asp$^{348}$ 및 His$^{374}$ 이 catalytic triad를 구성하고 있는 것으로 추정되었다. 여러 생물종의 EH서열을 기능적 및 구조적 domain 서열을 기초로 하여 multiple sequence alignment를 행하고 Neighbor-Joining/UPGMA method를 이용하여 계통수를 복원한 결과 다른 생물종들의 EH와의 진화거리는 서로 1.841∼2.682로 멀었으나 EH의 기능을 가지기 위한 oxyanion hole 및 a/$\beta$ hydrolase fold family의 catalytic triad는 잘 보존되고 있어 공통조상으로부터 진화되어 왔음을 알 수 있었다.

• Molecules and Cells
• /
• 제40권4호
• /
• pp.299-306
• /
• 2017

The transcriptional activator AphB has been implicated in acid resistance and pathogenesis in the food borne pathogens Vibrio vulnificus and Vibrio cholerae. To date, the full-length AphB crystal structure of V. cholerae has been determined and characterized by a tetrameric assembly of AphB consisting of a DNA binding domain and a regulatory domain (RD). Although acidic pH and low oxygen tension might be involved in the activation of AphB, it remains unknown which ligand or stimulus activates AphB at the molecular level. In this study, we determine the crystal structure of the AphB RD from V. vulnificus under aerobic conditions without modification at the conserved cysteine residue of the RD, even in the presence of the oxidizing agent cumene hydroperoxide. A cysteine to serine amino acid residue mutant RD protein further confirmed that the cysteine residue is not involved in sensing oxidative stress in vitro. Interestingly, an unidentified small molecule was observed in the inter-subdomain cavity in the RD when the crystal was incubated with cumene hydroperoxide molecules, suggesting a new ligand-binding site. In addition, we confirmed the role of AphB in acid tolerance by observing an aphB-dependent increase in cadC transcript level when V. vulnificus was exposed to acidic pH. Our study contributes to the understanding of the AphB molecular mechanism in the process of recognizing the host environment.

• BMB Reports
• /
• 제40권5호
• /
• pp.773-782
• /
• 2007

Different isolates of the soil bacterium Bacillus thuringiensis produce multiple crystal (Cry) proteins toxic to a variety of insects, nematodes and protozoans. These insecticidal Cry toxins are known to be active against specific insect orders, being harmless to mammals, birds, amphibians, and reptiles. Due to these characteristics, genes encoding several Cry toxins have been engineered in order to be expressed by a variety of crop plants to control insectpests. The cotton boll weevil, Anthonomus grandis, and the fall armyworm, Spodoptera frugiperda, are the major economically devastating pests of cotton crop in Brazil, causing severe losses, mainly due to their endophytic habit, which results in damages to the cotton boll and floral bud structures. A cry1Ia-type gene, designated cry1Ia12, was isolated and cloned from the Bt S811 strain. Nucleotide sequencing of the cry1Ia12 gene revealed an open reading frame of 2160 bp, encoding a protein of 719 amino acid residues in length, with a predicted molecular mass of 81 kDa. The amino acid sequence of Cry1Ia12 is 99% identical to the known Cry1Ia proteins and differs from them only in one or two amino acid residues positioned along the three domains involved in the insecticidal activity of the toxin. The recombinant Cry1Ia12 protein, corresponding to the cry1Ia12 gene expressed in Escherichia coli cells, showed moderate toxicity towards first instar larvae of both cotton boll weevil and fall armyworm. The highest concentration of the recombinant Cry1Ia12 tested to achieve the maximum toxicities against cotton boll weevil larvae and fall armyworm larvae were 230 ${\mu}g/mL$ and 5 ${\mu}g/mL$, respectively. The herein demonstrated insecticidal activity of the recombinant Cry1Ia12 toxin against cotton boll weevil and fall armyworm larvae opens promising perspectives for the genetic engineering of cotton crop resistant to both these devastating pests in Brazil.

• International Journal of Industrial Entomology and Biomaterials
• /
• 제3권1호
• /
• pp.19-23
• /
• 2001

Two B. thuringiensis strains, which possess mosquitocidal activities, were isolated from Korean soil samples and named K-1205-1 and K-1381-1. Serological studies indicated that K-1205-1 and K-1381-1 belonged to B. thuringiensis subsp. kurstaki (H3a3b3c) and subsp. aizawai (H7), respectively. K-1205-1 produced typical bipyramidal parasporal inclusions, but K-1381-1 produced irregular bipyramidal shape. Total plasmid DNA patterns analysis shewed that K-1205-1 and K- 1381-1 were different from their reference strains, subsp. kurstaki and subsp. aizawai, respectively, in high molecules, whereas their crystal protein patterns showed no difference. The cry gene contents of K-1205-1 and K-1381-1 were identical with those of the reference strains. Mosquitocidal activities of crystal proteins produced by K-1205-1 and K-1381-1 were significantly high by about 40-50 folds at $LC_50$ when compared to those of subsp. kurstaki and subsp. aizawai. Finally, in southern blot analysis using cry1A-type specific probe, K-1205-1 and K-1381-1 had different bands from subsp. kurstaki and subsp. aizawai, respectively. In conclusion, our results suggest that K-1205-1 and K-1381-1 appear to be new moquitocidal B. thuringiensis strains isolated from Korean soil.

• 한국잠사곤충학회지
• /
• 제35권2호
• /
• pp.120-128
• /
• 1993

B. t k HD-1 균주에 0.002% SDS와 0.5$\mu$g/ml EtBr을 처리한 결과, 9개의 cry- 변이균주를 분리하였으며 또한 B. t k HD-1균주와 B. cereus 569균주를 혼합배양하는 방법으로 mating 실험을 수행하여 B. t k HD-1으로부터 일부 plasmid가 전이된 11개의 cry+ B. cereus와 2개의 cry=B. cereus를 분리하고 plasmid수와 분자량을 조사하였다. B. t k HD-1의 경우 9개의 plasmid가 존재하였고 일부 plasmid가 curing된 B. t k HD-1변이균주의 경우 29Md plasmid나 44Md plasmid가 반드시 존재하였으나, cry- 변이균주에는 29Md 이상의 모든 plasmid가 소실되어 내독소 단백질 합성에 관여하는 유전자가 기억된 plamid를 결정하였다.

• 한국결정학회지
• /
• 제7권2호
• /
• pp.120-125
• /
• 1996

Salmonella typhimurium LT2에서 deoxy-thymidine diphosphate-D-gluxose-4,6-dehydratase의 유전자를 재조합한 Escherichia coli BL21 clone으로부터 dTDP-D-glucose dehydratase를 분리 정제한 후, 이 효소의 단결정을 상온에서 sitting drop 기체확산법으로 성장시켰다. 결정은 효소에 기질이 포함되어 있는 것과 포함되어 있지 않는 것 모두가 얻어지며, 이 때 침전제는 1.6-2.0 M Na, K 인산 완충용액(pH 8.0)을 사용하였다. 이 단결정은 최소 2.5Å의 분해능으로 회절하였으며, 공간군은 hexagonal한 P61이고, 격자의 크기는 a=b=168.54Å, c=81.08Å이었다. Asymmetric unit에는 이량체 한분자를 포함하고 있으며 단백질 질량당 결정의 부피는 VM=2.4Å3/Da, 용매의 함유율은 부피를 기준으로 64%였다.

• International Journal of Industrial Entomology and Biomaterials
• /
• 제11권1호
• /
• pp.57-61
• /
• 2005

A strain of Bacillus thuringiensis that showed signifi­cantly high toxicity to Plutella xylostella was isolated from a dust sample collected from Chinese tobacco warehouse and characterized. The isolate named B. thuringiensis LY-99 was determined to belong to subsp. alesti (H3a3c) by an H antisera agglutination test and produced bipyramidal inclusions. Plasmid and crystal protein patterns of the LY-99 were different from those of the reference strain, subsp. alesti. PCR analysis with specific primers revealed that this isolate contained abundant cry genes including crylAa, crylAc, crylB, crylD, crylE, crylF and cry2 genes, which was absolutely different from cry gene profile of the subsp. alesti. In addition, insecticidal activity of the LY-99 against P. xylostella larvae was about 44 times higher than that of the subsp. alesti.