• Title/Summary/Keyword: integrase

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Characterization of Human Foamy Virus Integrase Mutant (인간 포미바이러스 인테그라제 돌연변이의 특성)

  • Kang Seung Yi;Oh Soo A;Lee Hak Sung;Han Sung Tai;Shin Cha-Gyun
    • YAKHAK HOEJI
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    • v.49 no.3
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    • pp.198-204
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    • 2005
  • Human foamy virus (HFV) integrase mediates integration of viral c-DNA into cellular DNA. In this process, HFV integrase recognizes its own viral DNA specifically and catalyzes insertion of viral c-DNA. In order to study catalytic domains and residues, three deletion mutants and two point mutants of HFV integrase were constructed and analyzed with respect to enzymatic activities. The C-terminal deletion mutant showed decreased enzymatic activities while the N-terminal deletion mutant lost the activities completely, indicating that the N-terminal domain is more important than the C-terminal domain in enzymatic reaction. The point mutants, in which an aspartic acid at the 164th position or a glutamic acid at the 200th position of the HFV integrase protein was changed to an alanine, lost the enzymatic activities completely. However, they were well complemented with other defective deletion mutants to recover enzymatic activities partially. Therefore, these results suggest that the aspartic acid and glutamic acid at the respective 164th and 200th positions are catalytic residues for enzymatic reaction.

Multiple Functions of the Amino-terminal Domain of Bacteriophage Lambda Integrase: A New Member of Three-stranded $\beta-sheet$ DNA-binding Proteins

  • Cho Eun Hee
    • Proceedings of the Microbiological Society of Korea Conference
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    • 2002.10a
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    • pp.159-161
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    • 2002
  • Bacteriophage lambda integrase carries out the site-specific recombination of lambda. Integrase contains two DNA binding domains with distinct sequence specificity, namely arm-type binding and core-type binding domains. The amino-terminal arm-binding domain is structurally related to the three-stranded $\beta-sheet$ family of DNA-binding domains. Integrase binding to the high affinity arm-type site by the amino-terminal domain facilitates Int binding to the low affinity core-type site, where the cleavage and strand exchange occurs. The amino-terminal domain of Int also modulates the core-binding and catalysis through intramolecular domain-domain interaction and/or intermolecular interactions between Int monomers. In addition, the amino-terminal domain interacts cooperatively with excisionase during excision. This indicates that amino-terminal domain of Int plays an important role in formation of proper higher-order nucleoprotein structure required for lambda site-specific recombination.

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Phenotypes of Integrase-Mutated Human Immunodeficiency Virus Type-1(HIV-1)

  • ;Chris M. Farnet;William A. Haseltine
    • Proceedings of the Korean Society of Applied Pharmacology
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    • 1993.04a
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    • pp.92-92
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    • 1993
  • Point mutations in a highly conserved central region of the HIV-1 integrase protein were analyzed for their effects on viral replication and virion morphogenesis. Conservative amino acid replacements of two amino acid residues invariant un retroviral integrases, D116 and E152 of HIV-1, as well as the highly conserved amino acid S147, completely blocked viral replication in two CD4$\^$+/ human T cell lines. Mutation of four other highly conserved amino acids in the region had no detectable effect on viral replication, while Mutations at two positions, N117 and Y143, resulted in viruses with a delayed replication phenotype. Characteristic and reproducible defects id virion core structure were observed by electron microscopic analysis of sore of the replication defective integrase point mutants, indicating that mutant integrase proteins can interfere with the process of virion core maturation.

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Site-Specific Recombination by the Integrase MJ1 on Mammalian Cell (동물 세포 내에서 MJ1 인티그라제에 의한 부위 특이적 재조합)

  • Kim, Hye-Young;Yoon, Bo-Hyun;Chang, Hyo-Ihl
    • Microbiology and Biotechnology Letters
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    • v.39 no.4
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    • pp.337-344
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    • 2011
  • Integrase MJ1 from the bacteriophage ${\Phi}FC1$ carries out recombination between two DNA sequences (the phage attachment site, attP and the bacterial attachment site, attB) in NIH3T3 mouse cells. In this study, the integration vector containing attP, attB and the integrase gene MJ, was constructed. The integration mediated by integrase MJ1 in Escherichia coli led to excision of LacZ. Therefore, the frequency of integration was measured by the counting of the white colony, which is detectable on X-Gal plates. The extrachromosomal integration in NIH3T3 mouse cells was monitored by the expression of the green fluorescent protein (GFP) as a reporter. To demonstrate integration mediated integrase MJ1 in NIH3T3 cells, vectors containing attP and attB were co-transfected into NIH3T3 cells. The integration was confirmed by fluorescent microscopy. The expression of GFP was induced in NIH3T3 cells expressing MJ1 without accessory factors. By contrast, the excision mediated by the MJ1 between attR and attL had no effect on the expression of GFP. These results suggest that integrase MJ1 may enable a variety of genomic modifications for research and therapeutic purposes in higher living cells.

Explorations of the Electrostatic Character of a Model of Human Immunodeficiency Virus Type 1 Integrase to Offer a Prediction for the Orientation and Nature of DNA binding

  • Jung, Eun-Sun;Kwon, Yong-Jung
    • Journal of Industrial Technology
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    • v.26 no.B
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    • pp.163-171
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    • 2006
  • Human immunodeficiency virus type 1 (HIV-1) integrase plays a critical role in the life cycle of the HIV virus. An ability to accurately map its electrostatic potential, and then use this information to predict the manner in which DNA will bind to the active site of the catalytic domain could provide a foundation for inhibitory design. Attempts to discern the crystal structure of HIV-1 integrase have proven problematic, especially in the region of enzymatic activity, that being those residues involved in the catalysis of the integration of viral DNA into the host cell. However, there is a structural correlation in to the region of interest with avian sarcoma virus (ASV), so a homology model utilizing this similarity was constructed to approximate the behavior/structure of the undetermined portions of the HIV-1 integrase crystal. After this model was constructed and its energy minimized, electrostatic calculations were carried out on the substance, so that an electrostatic potential map was constructed. Using this information, it was determined that DNA binding was oriented so as to exploit the regions of positive potential nearby the active site, as well as the positive potential of the magnesium cofactors.

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HIV/AIDS Management: Dolutegravir Based Antiretroviral Drug Therapy

  • John, Ikpeama Osita;Emmanuel, Okoh Emeka;Anthonia, Ikpeama Chizoba;Joy, Ikpeama Chinwe;Adimabua, Okafor Patrick;Osazuwa, Igbineweka Osa;Andrew, Ikpeama Emeka;Mariam, Onuzulike Nonye;Gami, Hilary Tumba
    • The Korean Journal of Food & Health Convergence
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    • v.6 no.4
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    • pp.17-19
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    • 2020
  • HIV/AIDS disease still remain a global pandemic and it's management has undergone series of treatment changes and improvement although there is still no permanent cure.Dolutegravir belongs to a group of HIV drugs called integrase inhibitors. Integrase inhibitors block an HIV enzyme called integrase. By blocking integrase, integrase inhibitors prevent HIV from multiplying and can reduce the amount of HIV in the body.Dolutegravir combination based regimen has turned out to be very effective (antiviral) with negligible rare side effects on clients. This drug (Dolutegravir based regimen) combination has successfully increased the appetite for food of all the clients, unlike others and has shown to reduce viral load in the most shortest period ever. It can be deduced that development of resistant mutant virus will be reduced if not eliminated with dolutogravir based regimen.The role of Continuous adherence counseling has shown to improve clients treatment management. It is important to note that the availability of food has direct effect on the economic status or financial weight on the client. Hence the progress that is increase in body mass index (BMI) is a direct impact of the availability of food for the clients.

Biochemical characteristics of functional domains using feline foamy virus integrase mutants

  • Yoo, Gwi-Woong;Shin, Cha-Gyun
    • BMB Reports
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    • v.46 no.1
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    • pp.53-58
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    • 2013
  • We constructed deletion mutants and seven point mutants by polymerase chain reaction to investigate the specificity of feline foamy virus integrase functional domains. Complementation reactions were performed for three enzymatic activities such as 3'-end processing, strand transfer, and disintegration. The complementation reactions with deletion mutants showed several activities for 3'-end processing and strand transfer. The conserved central domain and the combination of the N-terminal or C-terminal domains increased disintegration activity significantly. In the complementation reactions between deletion and point mutants, the combination between D107V and deletion mutants revealed 3'-end processing activities, but the combination with others did not have any activity, including strand transfer activities. Disintegration activity increased evenly, except the combination with glutamic acid 200. These results suggest that an intact central domain mediates enzymatic activities but fails to show these activities in the absence of the N-terminal or C-terminal domains.

Characterization of Ty3-gypsy-like Elements in Hibiscus syriacus

  • Jeung, Ji-Ung;Cho, Sung Ki;Lee, Seung Jae;Shin, Jeong Sheop
    • Molecules and Cells
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    • v.19 no.3
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    • pp.318-327
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    • 2005
  • Southern blot analysis revealed a ubiquitous distribution and high copy number of Ty3-gypsy-like elements in the genome of Hibiscus syriacus. Comparative phylogenetic analysis of the large subunit of Rubisco and the integrase region of Ty3-gypsy elements in various plant species indicated that the retrotransposon-like sequences have different evolutionary histories and their own unique polymorphism in the H. syriacus population. Sequence-tagged site-restriction fragment length polymorphisms (STS-RFLP) analysis also indicated great variability in the numbers and sequences of Ty3-gypsy-like elements within H. syriacus varieties. Ty3-gypsy-like elements may still be active within H. syriacus, since Northern analysis of wounded leaves of H. syriacus variety Saehan with a probe for the integrase domain gave strong hybridization signals. The sequence heterogeneity and ubiquity of the Ty3-gypsy-like elements in H. syriacus genomes could provide reliable DNA markers for line identification as well for the analysis of genetic diversity in H. syriacus.

Analysis of Lambda Site-specific Recombination Inermediates Generated by Synthetic Substrates (합성 기질에 의해 형성된 Lambda Site-specific Recombination 중간 대사물의 분석)

  • 이나영;유승구
    • Microbiology and Biotechnology Letters
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    • v.23 no.3
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    • pp.282-287
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    • 1995
  • Integrase (Int) carries out the cutting and resealing of attachment (att) site DNA via a covalent Int-DNA intermediate. A family of synthetic substrate DNAs was designed to accumulate Int-DNA intermediate. Int-DNA intermediates accumulated by half substrate was analyzed by SDS- KCI precipitation and restriction digestion. The results showed that Int-half DNA intermediate was circular and contained covalently bound Int molecule. Int-DNA intermediates were also trapped with three other kinds of synthetic substrates.

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