• Title/Summary/Keyword: Protein-Coding Region

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Expression of the 38 kDa Protein of Mycobacterium tuberculosis in M. bovis BCG and Use in the Serodiagnosis of Tuberculosis

  • Cho, Sang-Nae;Kim, Hee-Jin;Lee, Hye-Young;Kim, Seung-Chul;Kim, Joo-Deuk
    • The Journal of the Korean Society for Microbiology
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    • v.34 no.6
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    • pp.555-559
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    • 1999
  • The 38 kDa protein of Mycobacterium tuberculosis, which was known previously as antigen 5, has been extensively used in the serodiagnosis of tuberculosis. In an attempt to develop and evaluate a serodiagnostic test using the antigen, we expressed the 38 kDa protein in BCG and its seroreactivity was compared to that expressed in Escherichia coli. The coding region of the 38 kDa protein was amplified by PCR, and the gene was cloned into a Mycobacterium-E. coli shuttle expression vector pYMC-his and pQE30 expression vector and expressed in BCG and E. coli, respectively. Both recombinant 38 kDa proteins showed strong seroreactivity against pooled serum from tuberculosis patients. There was no significant difference in seroreactivity between the two recombinant antigens in sera from the far advanced tuberculosis patients. However, of 25 tuberculosis patients graded as "minimal" by chest X-ray, 5 (20.0%) were seropositive by r38 kDa expressed in E. coli, while 8 (32.0%) by that expressed in BCG. Likewise, higher seroreactivity by r38 kDa expressed in BCG was found in sera from the moderately advanced tuberculosis. This study thus indicates that the recombinant 38 kDa expressed in BCG is more effective than that expressed in E. coli in detecting antibodies to the native 38 kDa protein of M. tuberculosis in sera from minimally affected tuberculosis patients.

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Nucleotide Sequence Analyses of p10 Gene and its Promoter of Hyphantria cunea Nuclear Polyhedrosis Virus (Hyphantria cunea Nuclear Polyhedrosis Virus p10유전자와 프로모터의 염기서열 결정)

  • Park, Sun-A;Cha, Sung-Chul;Chang, Jae-Hyeok;Lee, Hyung-Hoan
    • The Journal of Korean Society of Virology
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    • v.26 no.1
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    • pp.131-137
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    • 1996
  • The sequences of p10 gene its promoter of Hyphantria cunea NPV were determined. According to the sequence analysis, the putative p10 gene ORF has 285 bp. The 5'-non-coding leader sequence of the p10 gene promoter contained the TATA box and the putative transcription initiation site TAAG motif. Poly (A) tail signals, AATAAA sequence was at site 65 base upstream from the 3' terminus. The deduced amino acid sequence of p10 protein was 95 with a predicted molecular weight of 10.26 kDa. In the p10 protein sequence, a hydrophobic region was present at the N-terminus of the protein, whereas the C-terminus was highly hydrophilic. The p10 protein of H. cunea NPV did not contain cysteine, histidine, trytophan, tryptophane, tyrosine, glutamine and asparagine residues.

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Association of Tyrosinase (TYR) and Tyrosinase-related Protein 1 (TYRP1) with Melanic Plumage Color in Korean Quails (Coturnix coturnix)

  • Xu, Ying;Zhang, Xiao-Hui;Pang, You-Zhi
    • Asian-Australasian Journal of Animal Sciences
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    • v.26 no.11
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    • pp.1518-1522
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    • 2013
  • TYR (Tyrosinase) and TYRP1 (Tyrosinase-related protein 1) play crucial roles in determining the coat color of birds. In this paper, we aimed to characterize the relationship of TYR and TYRP1 genes with plumage colors in Korean quails. The SNPs were searched by cDNA sequencing and PCR-SSCP in three plumage color Korean quails (maroon, white and black plumage). Two SNPs ($367T{\rightarrow}C$ and $1153C{\rightarrow}T$) were found in the coding region of TYRP1 gene, but had no significant association with plumage phenotype in Korean quails. The expression of TYR was higher in black plumage quails than that in maroon plumage quails. In contrast, the expression of TYRP1 was lower in black plumage quails than that in maroon plumage quails. This study suggested that the melanic plumage color in Korean quails may be associated with either increased production of TYR or decreased production of TYRP1.

Isolation and Characterization of Chlorella Virus from Fresh Water in Korea and Application in Chlorella Transformation System

  • Park, Hye-Jin;Yoon, Hong-Mook;Jung, Heoy-Kyung;Choi, Tae-Jin
    • The Plant Pathology Journal
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    • v.21 no.1
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    • pp.13-20
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    • 2005
  • Chlorella viruses are large icosahedral, plaque-forming, dsDNA viruses that infect certain unicellular, chlorellalike green algae. The genomic DNA of over 300 kb contains many useful genes and promoters. Over 40 chlorella viruses have been isolated from fresh water in Korea since 1998. The viruses were amplified initially in chlorella strain NC64A, and pure isolates were obtained by repeated plaque isolation. SDS-PAGE analysis revealed similar but distinct protein patterns, both among the group of purified viruses and in comparison with the prototype chlorella virus PBCV-1. Digestions of the 330- to 350-kb genomic DNAs with 10 restriction enzymes revealed different restriction fragment patterns among the isolates. The tRNA-coding regions of 8 chlorella viruses were cloned and sequenced. These viruses contain 14-16 tRNA genes within a 1.2- to 2-kb region, except for the SS-1 isolate, which has a 1039-bp spacer in a cluster of 11 tRNA genes. Promoter regions of several early genes were isolated and their activities were analyzed in transformed chlorella. Some promoters showed stronger activity than commonly used CaMV 35S promoter and chlorella transformation vectors for heterologous protein are beings constructed using these promoters.

Characterization of a Glycoside Hydrolase Family 50 Thermostable β-agarase AgrA from Marine Bacteria Agarivorans sp. AG17

  • Nikapitiya, Chamilani;Oh, Chul-Hong;Lee, Young-Deuk;Lee, Suk-Kyoung;Whang, Il-Son;Lee, Je-Hee
    • Fisheries and Aquatic Sciences
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    • v.13 no.1
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    • pp.36-48
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    • 2010
  • An agar-degrading Agarivorans sp. AG17 strain was isolated from the red seaweed Grateloupia filicina collected from Jeju Island. A beta-agarase gene from Agarivorans sp. AG17 was cloned and designated as agrA. agrA has a 2,985 bp coding region encoding 995 amino acids and was classified into the glycoside hydrolase family (GHF)-50. Predicted molecular mass of the mature protein was 105 kDa. His-tagged agrA was overexpressed in Escherichia coli and purified as a fusion protein. The enzyme showed 158.8 unit/mg specific activity (optimum temperature at $65^{\circ}C$ and pH 5.5 in acetate buffer) with unique biochemical properties (high thermal and pH stabilities). Enzyme produced neoagarohexaose, neoagarotetraose and neoagarobiose by degrading agar, and hydrolyzed neoagaro-oligosaccharides were biologically active. Hence the purified enzyme has potential for use in industrial applications such as the development of cosmetics and pharmaceuticals.

A Novel Alternaria Species Isolated from Peucedanum japonicum in Korea

  • Deng, Jian Xin;Cho, Hye Sun;Paul, Narayan Chandra;Lee, Hyang Burm;Yu, Seung Hun
    • Mycobiology
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    • v.42 no.1
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    • pp.12-16
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    • 2014
  • We isolated and examined a new Alternaria sp., which causes leaf spots on Peucedanum japonicum in Korea, by using molecular and morphological methods. Phylogenetic analysis based on a combined internal transcribed spacer region analysis and two protein-coding genes (gpd and Alt a1) demonstrated that the causal fungus was most closely related to A. cinerariae and A. sonchi, and relevant to A. brassicae. However, conidial morphology indicated that it is a novel species within the genus Alternaria, and therefore we have assigned the fungus a new name in this study.

Conserved Regions in Mitochondrial Genome Sequences of Small Mammals in Korea

  • Kim, Hye Ri;Park, Yung Chul
    • Journal of Forest and Environmental Science
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    • v.28 no.4
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    • pp.278-281
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    • 2012
  • Comparative sequence analyses were conducted on complete mtDNA sequences from four small mammal species in Korea and revealed the presence of 30 well conserved sequences in various regions of the complete mtDNA sequences. The conserved sequences were found in 9 regions in protein coding genes, 10 regions in tRNA genes, 10 in rRNA genes, one region in replication origin and 2 regions in D loop. They could be used to design primers for amplifying complete mtDNA sequences of small mammals.

Isolation and Characterization of the C-type Lysozyme Gene from the Common Cutworm Spodoptera litura

  • Kim, Jong-Wan;Yoe, Sung-Moon
    • Animal cells and systems
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    • v.13 no.3
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    • pp.345-350
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    • 2009
  • We have isolated and characterized a new insect chicken type (c-type) lysozyme gene from the common cutworm, Spodoptera litura. The full-length cDNA of Spodoptera lysozyme is cloned by rapid amplification of cDNA ends PCR (RACE-PCR). The isolated cDNA consists of 1039 bp including the coding region for a 142-amino acid residue polypeptide, which included a signal peptide of 21-amino acid residue and a mature protein of 121-amino acid residue. The predicted molecular weight of mature lysozyme and its theoretical isoelectric point from amino acid composition is 13964.8 Da and 9.05, respectively. The deduced amino acid sequence of Spodoptera lysozyme gene shows the highest similarity (96.7%) to Spodoptera exigua lysozyme among other lepidopteran species. Amino acid sequence comparison with other the c-type lysozymes, Spodoptera lysozyme has the completely conserved $Glu^{32}$ and $Asp^{50}$ of the active site and eight Cys residues are completely conserved in the same position as that of other lepidopteran lysozymes.

Cloning, Sequencing and Expression of dTDP-D-Glucose 4,6-Dehydratase Gene from Streptomyces antibioticus $T\ddot{u}99$, a Producer of Chlorothricin

  • Sohng, Jae-Kyung;Yoo, Jin-Cheol
    • BMB Reports
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    • v.29 no.3
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    • pp.183-191
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    • 1996
  • DNA fragments, homologous to the dTDP-D-glucose 4,6-dehydratase gene, obtained from the genomic DNA of Streptomyces antibioticus $T\ddot{u}99$, a producer of the unusual macrolide antibiotic chlorothricin, were cloned and sequenced. This dehydratase gene was designated as oxil. The coding region of the oxil gene is composed of 987 bp, and analysis of the DNA sequence data reveals sequences for the gene products of 329 amino acids (molecular weight of 36,037). The deduced amino acids are 59% identical to the StrE, dTDP-D-glucose 4,6-dehydratase from the streptomycin pathway. The oxil's function was examined by expressing it in E. coli using the T7 RNA polymerase/promoter system (pRSET) to produce an active fusion protein including a his tag. This enzyme shows specificity of substrate, specific only to dTDP-D-glucose.

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Functional, Genetic, and Bioinformatic Characterization of Dextransucrase (DSRBCB4) Gene in Leuconostoc mesenteroides B-1299CB4

  • Kang, Hee-Kyoung;Kim, Young-Min;Kim, Do-Man
    • Journal of Microbiology and Biotechnology
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    • v.18 no.6
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    • pp.1050-1058
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    • 2008
  • A gene encoding a dextransucrase (dsrBCB4) that synthesizes only ${\alpha}$-1,6-linked dextran was cloned from Leuconostoc mesenteroides B-1299CB4. The coding region consisted of an open reading frame (ORF) of 4,395 bp that coded a 1,465-amino-acids protein with a molecular mass of 163,581 Da. The expressed recombinant DSRBCB4 (rDSRBCB4) synthesized oligosaccharides in the presence of maltose or isomaltose as an acceptor, plus the products included ${\alpha}$-1,6-linked glucosyl residues in addition to the maltosyl or isomaltosyl residue. Alignments of the amino acid sequence of DSRBCB4 with glucansucrases from Streptococcus and Leuconostoc identified conserved amino acid residues in the catalytic core that are critical for enzyme activity. The mutants D530N, E568Q, and D641N displayed a 98- to 10,000-fold reduction of total enzyme activity.