• Title/Summary/Keyword: Protein Mutation

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THE IMMUNOHISTOCHEMICAL STUDY ON THE EXPRESSION OF p53 PROTEIN AND CYCLIN D1 IN ORAL SQUAMOUS CELL CARCINOMAS (구강 편평세포암종에서 p53 단백과 Cyclin D1발현에 대한 면역조직화학적 연구)

  • Kim, Jong-Yub;Kim, Kyung-Wook;Lee, Jae-Hoon;Kim, Chang-Jin
    • Maxillofacial Plastic and Reconstructive Surgery
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    • v.21 no.2
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    • pp.139-148
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    • 1999
  • Oral cancer is a common neoplasm in humans and etiologic mechanism is not well known, so treatment and evaluation of oral cancer is difficult problem. Traditional TNM classification between prognosis of tumors and classification of histopathologic differentiation has problem like lack of objectivity through operators. In molecular biology, cancer is developed by alteration of activation of oncogene and/or inactivation of tumor suppressor gene. The p53 gene, one of the tumor suppresor genes, is believed to play an important role through mutation and overexpression in the progression of human cancers. The p53 mutation is most frequent genetic disorder in humans. The Cyclin D1 has tumor suppresion activity by regulation of cell cycle. The Cyclin D1 regulate activity of Rb tumor suppresor gene by stimulation of CDK4 The purpose of this study was to observe the expression of p53 protein and Cyclin D1 in oral squamous cell carcinoma, and to get expectation of the malignancy and prognosis of oral squamous cell carcinoma. Using the 15 cases of squamous cell carcinoma and the microscopic H&E and immunohistochemical stain. We divided it into 3 groups according to the stain extent, clinical stage and histologic differentiation. The results were as follows1.In the features of immunohistochemical stain of 15 cases of squamous cell carcinoma, positive reaction of p53 was identified in 8 cases (53.3%) and positive reaction of cyclin D1 was identified in 3 cases (20%). Both positive reaction of p53 protein and Cyclin D1 was show in only one case. 2.8 of p53 positive cases were linked in 87.5% of the end stage tumor, 62.5% of neck node involvement, 87.5% of poorly and moderately histopathplogic differentiation. 3. All 3 of Cyclin D1 positive cases were linked in the end stage tumor, neck node involvement, poorly and moderately histopathologic differentiation. From above results, expression of p53 protein was identified in 53.3% of 15 cases and these results mean oral squamous cell carcinoma was drived by mutation of p53 protein. Especially, highly positive reaction of p53 protein and Cyclin D1 was identified in cases that involvement of neck lymph node and the end stage tumors and it means that the evaluation of p53 protein and Cyclin D1 was useful for evaluation of malignant tumor as specific tumor marker.

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DNA Microarray and Gene Ontology Enrichment Analysis Reveals That a Mutation in opsX Affects Virulence and Chemotaxis in Xanthomonas oryzae pv. oryzae

  • Kim, Hong-Il;Park, Young-Jin
    • The Plant Pathology Journal
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    • v.32 no.3
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    • pp.190-200
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    • 2016
  • Xanthomonas oryzae pv. oryzae (Xoo) causes bacterial leaf blight (BLB) in rice (Oryza sativa L.). In this study, we investigated the effect of a mutation in opsX (XOO1056), which encodes a saccharide biosynthesis regulatory protein, on the virulence and bacterial chemotaxis of Xoo. We performed DNA micro-array analysis, which showed that 63 of 2,678 genes, including genes related to bacterial motility (flagellar and chemotaxis proteins) were significantly downregulated ($<\;-2\;log_2$ fold changes) by the mutation in opsX. Indeed, motility assays showed that the mutant strain was nonmotile on semisolid agar swarm plates. In addition, a mutant strain (opsX::Tn5) showed decreased virulence against the susceptible rice cultivar, IR24. Quantitative real-time RT-PCR reaction was performed to confirm the expression levels of these genes, including those related to flagella and chemotaxis, in the opsX mutant. Our findings revealed that mutation of opsX affects both virulence and bacterial motility. These results will help to improve our understanding of Xoo and provide insight into Xoo-rice interactions.

Frequency and Type of Disputed rpoB Mutations in Mycobacterium tuberculosis Isolates from South Korea

  • Jo, Kyung-Wook;Lee, Soyeon;Kang, Mi Ran;Sung, Heungsup;Kim, Mi-Na;Shim, Tae Sun
    • Tuberculosis and Respiratory Diseases
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    • v.80 no.3
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    • pp.270-276
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    • 2017
  • Background: A disputed rpoB mutation is a specific type of rpoB mutation that can cause low-level resistances to rifampin (RIF). Here, we aimed to assess the frequency and types of disputed rpoB mutations in Mycobacterium tuberculosis isolates from South Korea. Methods: Between August 2009 and December 2015, 130 patients exhibited RIF resistance on the MTBDRplus assay at Asan Medical Center. Among these cases, we identified the strains with disputed rpoB mutation by rpoB sequencing analysis, as well as among the M. tuberculosis strains from the International Tuberculosis Research Center (ITRC). Results: Among our cases, disputed rpoB mutations led to RIF resistance in at least 6.9% (9/130) of the strains that also exhibited RIF resistance on the MTBDRplus assay. Moreover, at the ITRC, sequencing of the rpoB gene of 170 strains with the rpoB mutation indicated that 23 strains (13.5%) had the disputed mutations. By combining the findings from the 32 strains from our center and the ITRC, we identified the type of disputed rpoB mutation as follows: CTG511CCG (L511P, n=8), GAC516TAC (D516Y, n=8), CTG533CCG (L533P, n=8), CAC526CTC (H526L, n=4), CAC526AAC (H526N, n=3), and ATG515GTG (M515V, n=1). Conclusion: Disputed rpoB mutations do not seem to be rare among the strains exhibiting RIF resistance in South Korea.

Site-specific and deletional mutagenesis for two regions of Verotoxin-2 A gene encoding enzymatically active domain (Verotoxin-2 A 유전자의 효소활성 부위에 대한 위치특이적 변이 및 결손변이유발)

  • Kim, Yong-hwan;Kim, Sang-hyun;Cha, In-ho;Kim, Kyoung-shook;Lee, Young-choon
    • Korean Journal of Veterinary Research
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    • v.37 no.3
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    • pp.541-546
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    • 1997
  • There are two conserved regions with a significantly high amino acid sequence homology among the A subunits of STX, SLTs and ricin. To produce an inactive Verotoxin-2 (VT-2), two different mutants, pE167D and pDE5A, were constructed by site-directed mutagenesis, respectively, on the basis of the previous reports that two regions lie within the active-site clefts of the A subunits of ricin and STX family. The cytotoxicity ($10^3$ $CD_{50}/ml$) of VT-2 holotoxin with E167D mutation was reduced by $10^3$-fold compared with wild-type level. In addition, VT-2 with DE5A ($Trp_{202}GlyArgIleSer_{206}$) deletion mutation showed a significantly low cytotoxicity ($10^1$ $CD_{50}/ml$), resulting in $10^5$- and $10^2$-fold reductions, respectively, compared with the wild-type and E167D mutatant. SDS-PAGE for protein samples showed a 33-kDa band corresponding to the A subunit of VT-2. These results indicate that reduction in cytotoxic activity was affected not by amount of VT-2 protein produced but by mutation.

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Induction of Deletion Mutation for the Enzymatic Domain in the Shigatoxin2e A Subunit Gene of Esherichila coli O139 Isolates and Expression of Mutated Protein (분리 대장균 O139의 Shigatoxin2e A 유전자의 효소 활성부에 대한 결손변이 유발 및 변이 단백질의 발현)

  • Cho Eun-jung;Kim Do-kyong;Kim Sang-hyun;Kim Yeong-il;Lee Chul-hyun;Lee Woo-won;Son Won-geun;Shin Jong-Uk;Kim Yong-hwan
    • Journal of Veterinary Clinics
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    • v.22 no.4
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    • pp.386-391
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    • 2005
  • This study was done to produce a mutated protein inactivated cytotoxicity of Shigatoxin 2e (Stx2e) of E.coli O139 isolates by deletional mutagenesis of Stx2e A subunit gene encoding active-site cleft of enzymatic domain in ST2e holotoxin. Cytotoxicity of the toxoid expressed from the mutant Stx2e gene was compared with wild type Stx2e for development of vaccine candidate. A recombinant plasmid pED18 containing Stx2e gene ot E.coli O139 isolates was used to generate mutation plasmid. Deletion mutagenesis was conducted for Stx2e A subunit gene encoding enzymatically active domain by polymerase chain reaction (PCR) using ot designed primer to induce deletional mutation. DNA sequence analysis was confirmed that the pentamer (Typ 202- Ser 206) that lies within the proposed active-site cleft in the second region was completely deleted. A DNA fragment of 1.1 kb that encode the new mutant Stx2eA gene was inserted into plasmid pRSET vector digested with EcoRV-Hind III and named pEDSET The PEDSET was transformed in E. coli for expression of mutant protein and the protein was confirmed by SDS-PACE and Western-blotting. The protein expressed by the mutant was tested to confirm the reduction of cytotoxic activities on Vero cell using microcytotoxicity assay compared with wild type Stx2e, the cytotoxicity of deletional mutant protein was at least reduced by 3,000-fold on Vero cell.

SITE-DIRECTED MUTATION STUDY ON HYPERTHERMOSTABILITY OF RUBREDOXIN FROM PYROCOCCUS FURIOSUS USING MOLECULAR DYNAMICS SIMULATIONS IN WATER

  • Jung, Dong-Hyun;Kang, Nam-Sook;Jhon, Mu-Shik
    • Proceedings of the Korean Biophysical Society Conference
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    • 1996.07a
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    • pp.21-21
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    • 1996
  • The hyperthermostable protein, rubredoxin from Pyrococcus furiosus is 53-residue protein with a three-stranded anti-parallel $\beta$-sheet and several loops. To investigate the effect of changes of electrostatic and hydrophobic interactions on the structure and dynamic property of P. furiosus rubredoxin, molecular dynamics simulations in water were performed on three mesophilic rubredoxins, P, furiosus rubresoxin, and 5 mutants of P. furiosus rubredoxin. (omitted)

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EXAMINATION OF TYR-264 FOR ATPase ACTIVE SITE IN E.coli RecA PROTEIN BY SITE-DIRECTED MUTAGENESIS

  • Kwon, Yong-Kook;Bae, Jun-Seong;Hahn, Tae-Ryong
    • Journal of Photoscience
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    • v.2 no.1
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    • pp.27-29
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    • 1995
  • Site directed mutagenesis has been introduced to determine active site(s) and molecular structure of E. coli RecA protein. Recombinant DNAs were constructed by point mutation of Tyr-264 to Phe which assumed active site for binding and hydrolysis of ATP. RecA proteins were purified from recombinants containing wild type and mutant genes and analyzed for ATPase activity assay. Result suggests that Tyr-264 is involved in ATP binding rather than ATP hydrolysis.

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