• Journal of Life Science
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• v.15 no.2 s.69
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• pp.192-196
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• 2005

The SNF2/SW12 family comprises proteins from a variety of species with in vivo functions, such as transcriptional regulation, maintenance of chromosome stability during mitosis, and various types of DNA repair. This study was shown the characterization of hrp2+ gene which was isolated by PCR amplification using the conserved domain of SNF2 motifs. Sequence analysis of hrp2+ gene showed striking evolutionary conservation among the SNF2 family of proteins. The transcript of hrp2+ gene was found to be a 4.7 kb as identified by Northern hybridization. To investigate the inducibility of hrp2+ gene, transcript levels were examined after treating the cells to various DNA damaging agents. The transcripts of hrp2+ were induced by UV-irradiation. But the transcripts were not induced by treatment of $ 0.25\%$ Methylmethane sulfonate (MMS). These results implied that the effects of damaging agents are complex and different regulatory pathways exist for the induction of this gene. Hrp2 protein was purified near homogeneity by combination of affinity chromatography. We tested the purified Hrp2 protein for the helicase activity in an oligonucleotide release assay. However we were unable to detect any helicase activity associated with the Hrp2 protein, indicating that the helicase motifs in Hrp2 are merely indicators of a broader DNA-dependent ATPase activity.

• Animal cells and systems
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• v.6 no.4
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• pp.311-315
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• 2002

The RAD4 gene is essential for nucleotide excision repair in Saccharomyces cerevisiae. It has been known that the deduced amino acid sequence of Rad4 protein contains three DNA-dependent ATPase/helicase motifs. To determine the biochemical activities and functional role of RAD4 the Rad4 protein was expressed and purified. Immunoblot analysis showed a specific band of 21 kDa, which was well-matched with the size of open reading frame of the RAD4 gene. The purified Rad4 protein had no detectable helicase activity. However, the protein could interact with double stranded oligonucleotides, as judged by mobility shift assay. This result suggests that the Rad4 protein is a DNA binding protein.

• Environmental Mutagens and Carcinogens
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• v.16 no.2
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• pp.77-82
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• 1996

The RAD3 gene of Saccharomyces cerevisiae is required for excision repair and is essential for cell viability. RAD3 encoded protein possesses a single stranded DNA-dependent ATPase and DNA-RNA helicase activies. To examine the extent of conservation of structure and function of RAD3 during eukaryotic evolution, we have cloned the RAD3 homolog, HRD3, from the distantly related yeast Schizosaccharomyces pombe. Here, we report the partial cloning and characterization of HRD3 gene (Homologous of RAD3 gene) which was isolated by PCR amplification using conserved domain of Saccharomyces cerevisiae RAD3 gene. Chromosomal DNA isolated from S. pombe had similar restriction patterns to those from S. cerevisiae, as determined by Southern blot analysis. The 2. 8 kb transcript of mRNA was identified by Northern hybridization. The level of transcript did not increase upon UV-irradiation, suggesting that the HRD3 gene in S. pombe is not UV-inducible.

• The Journal of Korean Society of Virology
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• v.27 no.1
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• pp.29-37
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• 1997

To use recombinant viruses with wider host range as viral insecticides, we investigated the characteristics and pathogenicity of host range expanded recombinant viruses in insect cells. We compared host range expanded recombinant viruses, RecS-B6 and RecB-8, constructed by cotransfection of Autographa californica nuclear polyhedrosis virus (AcNPV) and Bombyx mori NPV (BmNPV), to host range expanded AcNPV, Ac-BH, by substitution of the 0.6 Kb fragment of the BmNPV helicase gene. Restriction endonuclease profiles of RecS-B6 and RecB-8 DNAs were different from those of parent viruses. Nucleotide sequence analysis of the 0.6 Kb region in the putative helicase gene of RecS-B6 and RecB-8 showed that their structures were identical to the counterpart region of BmNPV. Comparison of viral replication of these recombinant viruses in Sf-21 and BmN-4 cells showed that Ac-BH, compared to wild type viruses, replicated well in BmN-4 cells but poorly in Sf-21 cells. In contrast, RecS-B6 and RecB-8 replicated relatively well in both cells compared to parent viruses. These results may imply that random genomic recombinant viruses, RecS-B6 and RecB-8, possess better potential as viral pesticides than helicase-mediated recombinant virus, Ac-BH.

• Journal of Microbiology and Biotechnology
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• v.25 no.4
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• pp.492-495
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• 2015

This study provides evidence that RecG regulates the expression of the OxyR-independent gene mexA in Pseudomonas aeruginosa PAO1. A reduction in mexA expression was observed in the absence of RecG, but not OxyR, by northern blot and quantitative real-time PCR analyses. The canonical palindromic RecG binding sequence was present upstream of the mexA promoter, and bound purified RecG and single strand-binding protein. These data reveal a novel mechanism of OxyR-independent gene transcription by RecG.

• Journal of Life Science
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• v.6 no.3
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• pp.185-192
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• 1996

Bacteriophage T7 gene 2.5 protein, a single-stranded DNA binding protein, is required for T7 DNA replication, recombination, and repair. T7 gene 2.5 protein has two distinctive domains, DNA binding and C-terminal domain, directly involved in protein-protein interaction. Gene 2.5 protein participates in the DNA replication of Bacteriophage T7, which makes this protein essential for the T7 growth and DNA replication. What gene 2.5 protein makes important at T7 growth and DNA replication is its binding affinity to single-stranded DNA and the protein-protein important at T7 DNA replication proteins which are essential for the T7 DNA synthesis. We have constructed pGST2.5(WT) encoding the wild-type gene 2.5 protein and pGST2.5$\Delta $21C lacking C-terminal 21 amino acid residues. The purified GST-fusion proteins, GST2.5(WT) and GST2.5(WT)$\Delta$21C, were used for whether the carboxyl-terminal domain participates in the protein-protein interactions or not. GST2.5(WT) and GST2.5$\Delta$21C showed the difference in the protein-protein interaction. GST2.5(WT) interacted with T7 DNA polymerase and gene 4 protein, but GST2.5$\Delta$21C did not interact with either protein. Secondly, GST2.5(WT) interacts with gene 4 proteins (helicase/primase) but not GST2.5$\Delta$21C. these results proved the involvement of the carboxyl-terminal domain of gene 2.5 protein in the protein-protein interaction. We clearly conclude that carboxy-terminal domain of gene 2.5 protein is firmly involved in protein-protein interactions in T7 replication proteins.

• Journal of Sericultural and Entomological Science
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• v.38 no.1
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• pp.42-47
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• 1996

To investigate the host range determining factors of nuclear polyhedrois virus (NPV), Autographa california NPV and Bombyx mori NPV were coinfected into the two different cell lines, BmN-4 and Sf-9. The recombinant baculoviruses, RecS-A6 and RecB-727 which have an expanded host range, were isolated from Sf-9 and BmN-4 cell lines, respectively. The molecular biological characteristics of the recombinant baculoviruses were investigated. The pathogenicity of RecB-727 was similar to that of wild type BmNPV, while the pathogenicity of RecS-A6 was relatively lower than that of wild type BmNPV. The restriction enzyme digestion patterns of parental viruses and recombinant viruses showed that the recombinant virus has an expanded host range by genetic recombination. Southern blot analysis revealed that the p10 gene of RecB-727 was derived from AcNPV genomic DNA, while RecS-A6 has p10 gene of BmNPV in a viral genome. To investigate the host range expansion mechanism of recombinant baculovirus, HindIII-SacI 0.6 kb DNA fragments of RecS-A6 and RecB-727 were cloned and sequenced. The results showed that of wild type BmNPV helicase gene, suggesting that the expanded host range of recombinant baculoviruses was due to the insertion of BmNPV helicase gene into AcNPV viral genome.

• Korean Journal of Veterinary Research
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• v.58 no.1
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• pp.45-49
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• 2018

The helicase genes and hypervariable regions (HVRs) of three avian hepatitis E viruses (HEVs) detected at three different farms were sequenced and characterized. Two isolates (DW-L and GI-B2) were classified as genotype 2 and one isolate (GR-B) was classified as genotype 1. A phylogenetic tree, based on the helicase gene and HVR nucleotide sequences, revealed the newly detected viruses and other avian HEVs were classified similarly. Unlike previously reported avian HEVs, the DW-L isolate detected in broiler breeders with characteristic lesions of avian HEV had no proline-rich motif in its HVR, suggesting that the proline-rich motif is non-essential for viral replication and infection.

• Korean Journal of Environmental Biology
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• v.38 no.3
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• pp.350-354
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• 2020

Many parrots are considered endangered species due to threats from human activities. Gender determination is of great importance for biological studies and the conservation of endangered parrots. However, like other birds, gender determination in parrots is hindered due to the lack of external dimorphism between males and females. A molecular approach using the chromo-helicase-DNA binding protein 1 (CHD1) gene is commonly used for sexing birds. This study aimed to determine the gender of parrots from Korean zoos based on amplification and visualization of the partial CHD1 gene. The samples of 13 parrot species were collected from three different zoos in Korea and the extracted DNA templates were amplified using CHD1 gene primers. The gender of 27 samples of 13 species was determined by visualizing the PCR products on an agarose gel. While male parrots were indicated by a single band, female parrots were indicated by double bands. The findings provide additional information, which might be helpful for the management and care of parrots in Korean zoos.

• Journal of Life Science
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• v.11 no.2
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• pp.111-115
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• 2001

This study was designed to clone the SNF2/SW12 helicase-related genes from the fission yeast Schizosaccha-romyces pombe and thereafter to elucidate the common functions of the proteins in this family. The $hrp^{2+}$gene was cloned by polymerase chain reaction amplification using degenerative primers from conserved SNF2 motifs within the ERCC6 gene, which encodes a protein involved in DNA excision repair. Like other SNF2/SW12 family proteins, the deduced amino acid sequence of Hrp2 contains DNA-dependent ATPase/7 helicase domains as well as the chromodomain and the DNA binding domain. This configuration is similar to that of mCHD1 (mouse chromo-ATPase/helicase-DNA-dinding protein 1), suggesting that Hrp2 is a S. pombe homolog of mCHD1, which is thought to function in altering the chromatin structure to control the gene expression. To characterize the function of Hrp2, 4 Uracil-Hrp2 fusion protein, it was purified near homogeneity by affinity chromatography on $Ni^{2+}$-NTA agarose, DEAE-Sepharose ion exchange arid Sephacryl S-200 gel filtration chromatographies. The purified fusion protein exhibited DNA-dependent ATPase activity, which was stimulated by both double-stranded and single-stranded DNA. To determine the steady-state level of $hrp^{2+}$ transcripts during growth, cells were cultured in medium and collected at every 2hr to prepare total RNAs. The northern blot analysis showed that the level of $hrp^{2+}$ transcripts reached its maximum before the cells entered the exponential growth phase and then decreased gradually, This result implies that Hrp2 may be required at early stages of cell growth.h.