• International Journal of Industrial Entomology and Biomaterials
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• 제9권1호
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• pp.149-153
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• 2004

Here we report the molecular cloning of a LIM protein cDNA of the CRP (cysteine-rich protein) family from the mulberry longicorn beetle, Apriona, geramri. The A. germari LIM protein cDNA contains an open reading frame of 276 bp encoding 92 amino acid residues with a calculated molecular weight of approximately 10 kDa. The A. germari LIM protein contains the cysteine-rich consensus sequence of LIM domain and the glycine-rich consensus sequence observed in cysteine-rich protein family 1 (CRP1). The potential nuclear targeting signal is retained. The deduced amino acid sequence of the A. germari LIM protein cDNA showed 81 % identity to both Bombyx mori muscle LIM protein (Mlp) and Drosophila melanogaster Mlp60A and 77% to Epiblema scudderiana Mlp. Northern blot analysis showed that A. germari LIM protein is highly expressed in epidermis and muscle, and less strongly in midgut, but not in the fat body.

• International Journal of Industrial Entomology and Biomaterials
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• 제11권1호
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• pp.67-70
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• 2005

A cuticle protein gene, PbLCP12.1, from the white­spotted flower chafer, Protaetia brevitarsis, was isolated and characterized. The gene contains an ORF of 336 nucleotides capable of encoding a 113 amino acid polypeptide with a predicted molecular mass of 12,138 Da and pI of 4.15. The PbLCP12.1 protein contained a type-specific consensus sequence identifiable in other insect cuticle proteins. The deduced amino acid sequence of the PbLCP12.1 cDNA is most similar to Bombyx mori cuticle protein BmLCP18 (37$\%$ protein sequence identity). Northern blot analysis revealed that PbLCP12.1 showed the epidermis-specific expression.

• International Journal of Industrial Entomology and Biomaterials
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• 제11권1호
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• pp.71-74
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• 2005

A LIM protein gene homologue of the CRP (cysteine­rich protein) family in the whiter-spotted flower chafer, Protaetia brevitarsis, was cloned. The P. brevitarsis LIM protein cDNA encodes a 92 amino acid polypep­tide with a predicted molecular mass of 10,030 Da and a pI of 8.57. The P. brevitarsis LIM protein contains the cysteine-rich consensus sequence of LIM domain and the glycine-rich consensus sequence observed in the cysteine-rich protein family 1 (CRPl). The potential nuclear targeting signal is retained. The deduced amino acid sequence of the P. brevitarsis LIM protein cDNA showed 92$\%$ identity to another beetle, Apriona germari LIM protein. Northern blot analysis showed that P. brevitarsis LIM protein is highly expressed in epidermis and midgut, but not in the fat body.

• International Journal of Industrial Entomology and Biomaterials
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• 제6권2호
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• pp.157-162
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• 2003

The glutathione S-transferase (GSTs) are enzymes responsible for the protection of cells from chemical toxicants and oxidative stress. We describe here the cDNA sequence and mRNA expression of a putative GST from the mole cricket, Gryllotalpa orientalis. The G. orientalis GST cDNA sequences comprised of 621 bp encoding 207 amino acid residues. The multiple sequence alignment of G. orientalis GST gene with other known insect GSTs showed several conserved residues that may be essential for the enzymatic activity of the protein. Phylogenetic analysis of the deduced amino acid sequences of G. orientalis GST gene with other insect GST sequences revealed that the G. orientalis GST gene belongs to class I GST, forming a strong monophyletic group (100% bootstrap value) exclusively for class I GSTs from a diverse insect species. Northern blot analysis confirmed midgut-specific expression at transcriptional level, evidencing the midgut as a site for GST synthesis.

• 한국작물학회지
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• 제50권4호
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• pp.286-290
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• 2005

Ascorbate peroxidase (APX) plays a crucial role in the detoxification of hydrogen peroxide. APX activity is maintained significantly higher in the paraquat­treated leaves of the paraquat-tolerant Rehmannia glutinos. This study was conducted to understand structural and regulatory characteristics of APX gene in R. glutinosa. A putative APX cDNA clone (RgAPX1) was isolated from a leaf cDNA library using a partially sequenced expressed sequence tag clone. RgAPX1 is consisted of 1148 bp nucleotides and contains an open reading frame encoding a 250 amino acid-long polypeptide. Deduced RgAPX1 amino acid sequence shares higher sequence similarity to cytosolic APXs. RgAPX1. expression was higher in the leaf than in the flower and root. Southern blot result indicates the presence of one or two RgAPX1-related genes in R. glutinosa genome. RgAPX1 transcription was affected differentially by various stresses and phytohormone. The results indicate that RgAPXl is constitutively expressed in most tissues and its expression is modulated for the immediate and efficient detoxification of $H_2O_2$ under normal and stress conditions.

• Journal of Microbiology and Biotechnology
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• 제10권4호
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• pp.559-563
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• 2000

A dextransucrase gene (dsrB742) that expresses a dextransucrase to synthesize mostly ${\alpha}-1{\rightarrow}6$ linked dextran with a low amount (3-5%) of ${\alpha}-1{\rightarrow}3$ branching was cloned and sequenced from Leuconostoc mesenteroides B-742CB. The 6.1-kb PstI fragments were ligated with pGEM-3Zf(-) and transformed into E. coli $DH5{\alpha}$. The recombinant clone (pDSRB742) synthesized dextran on an agar plate containing 2% (w/v) sucrose. The dextran synthesized was hydrolyzed with Penicillium endo-dextranase. The hydrolyzate was composed of glucose, isomaltose, isomaltotriose, and branced pentasaccharide. The nucleotide sequence of dsrB742 showed one open reading frame (ORF) composed of 4,524 bp encoding dextrasnsucrase. The deduced amino acid sequence revealed a calculated molecular mass of 168.6 kDa. It also showed an activity band of 184 kKa on a non-denaturing SDS-PAGE (10%). The amino acid sequence of DSRB742 exhibited a 50% similarity with DSRA from L. mesenteroides B-1299, a 70% similarity with DSRS from L. mesenteroides B-512 (F, FMCM) and a 45-56% similarity with Streptococcal GTFs.

• Journal of Microbiology and Biotechnology
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• 제8권1호
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• pp.28-33
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• 1998

A gene coding for ${\beta}$-xylosidase from thermophilic xylanolytic Bacillus sp. KK-1 was cloned into Escherichia coli using plasmid pBR322. Recombinant plasmid DNAs were isloated from E. coli clones which were capable of hydrolyzing 4-methylumbelliferyl-${\beta}$-D xylopyranoside. Restriction analysis showed the DNAs to share a common insert DNA. Xylo-oligosaccharides, including xylotriose, xylotetraose, xylopentaose, and xylobiose were hydrolyzed to form xylose as an end product by cell-free extracts of the E. coli clones, confirming that the cloned gene from strain KK-1 is ${\beta}$-xylosidase gene. The ${\beta}$-xylosidase gene of strain KK-1 designated as xylB was completely sequenced. The xylB gene consisted of an open reading frame of 1,602 nucleotides encoding a polypeptide of 533 amino acid residues, and a TGA stop codon. The 3' flanking region contained one stem-loop structure which may be involved in transcriptional termination. The deduced amino acid sequence of the KK-1 ${\beta}$-xylosidase was highly homologous to the ${\beta}$-xylosidases of Bacillus subtilis and Bacillus pumilus, but it showed no similarity to a thermostable ${\beta}$-xylosidase from Bacillus stearothermophilus.

• BMB Reports
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• 제34권4호
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• pp.371-378
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• 2001

The Glycoprotein D (gD) gene of the HSV-1 strain F was cloned, sequenced, recombinated into the HcNPV (Hyphantria cunea nuclear polyhedrosis virus) expression vector and expressed in insect cells. The gD gene was located in the 6.43 kb BamHI fragment of the strainF. The open reading frame (ORF) of the gD gene was 1,185 by and codes 394 amino acid residues. Recombinant baculoviruses, GD-HcNPVs, expressing the gD protein were constructed. Spodoptera frugiperda cells, infected with the recombinant virus, synthesized a matured gX-gD fusion protein with an approximate molecular weight of 54 kDa and secreted the gD proteins into the culture media by an immunoprecipitation assay The fusion gD protein was localized on the membrane of the insect cells, seen by using an immunofluorescence assay The deduced amino acid sequence presents additional characteristics compatible with the structure of a viral glycoprotein: signal peptide, putative glycosylation sites and a long C-terminal transmembrane sequence. These results indicate the utility of the HcNPV-insect cell system for producing and characterizing eukaryotic proteins.

• 한국어병학회지
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• 제22권3호
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• pp.343-352
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• 2009

Natural-killer-cell-enhancing factor (NKEF) belongs to the newly defined peroxiredoxin (Prx) family. It was originally isolated from human erythroid cells. The black rockfish NKEF cDNA was identified through the expressed sequence tag (EST) analysis of PBLs libraries. The full-length NKEF cDNA was 1433 bp long and contained an open reading frame (ORF) of 594 bp that encoded 198 amino-acid residues. The 5' UTR had a length of 39 bp, and the 3’UTR 800 bp. The deduced amino-acid sequence of the black rockfish had a density 93.4, 92.9, 87.8, 85.8, 84.8, 83.8, 80.3, 79.7, 77.2, and 75.2% that of the pufferfish, olive flounder, channel catfish, zebrafish, chicken, common carp, Myotis lucifugus, cattle, human PrxI, rat PrxI, human NKEF-A, and Xenopus tropicalis, respectively. The NKEF gene was expressed in all the tissues of the black rockfish. The RT-PCR indicated that the NKEF transcripts were predominantly in the spleen and gill, less dominantly in the PBLs, head kidney, trunk kidney, and liver, and least in the intestine and muscles. This is the first report on the existence of the NKEF-A gene in black rockfish.

• Journal of Microbiology and Biotechnology
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• 제22권5호
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• pp.642-648
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• 2012

We determined the nucleotide sequence of the URA3 gene encoding orotidine-5'-phosphate decarboxylase (OMPDCase) of the erythritol-producing osmotolerant yeast Candida magnoliae by degenerate polymerase chain reaction and genome walking. Sequence analysis revealed the presence of an uninterrupted open-reading frame of 795 bp, encoding a 264 amino acid residue protein with the highest identity to the OMPDCase of the yeast Kluyveromyces marxianus. Phylogenetic analysis of the deduced amino acid sequence revealed that it shared a high degree of identity with other yeast OMPDCase homologs. The cloned URA3 gene successfully complemented the ura3 null mutation in Saccharomyces cerevisiae, revealing that it encodes a functional OMPDCase in C. magnoliae. An enzyme activity assay and reverse transcription polymerase chain reaction indicated that the expression level of the C. magnoliae URA3 gene in S. cerevisiae was not as high as that of the S. cerevisiae URA3 gene. The GenBank accession number for C. magnoliae URA3 is JF521441.