• BMB Reports
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• 제33권1호
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• pp.92-95
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• 2000

The HIV-1 p24(202-221) sequence ETINNEEEWDRVHPV HAGP contains a B-cell epitope with the earliest immune response and the highest antibody titer against anti-mouse sera obtained by immunization with p24 antigens. A novel mouse monoclonal antibody (mAb) was generated against the immunodominant B-cell epitope of the HIV-1 p24 capsid protein, p24(202-221). BALB/c mice were immunized with the four branched multiple antigenic peptide (MAP) containing the HIV-1p24(202-221) sequence, and antibody-secreting hybridoma were produced by fusion of mouse splenocytes with P3X63Ag8.653, mouse myeloma cells. One clone which produced the antigen-specific mAb named KI-24 (Isotype IgG1, light chain: ${\kappa}$) was identified. mAb KI-24 was highly specific for both the p24(202-221) and p24 proteins when analyzed by ELISA and Western blotting. Since p24(202-221) also contains a cytotoxic T-lymphocyte epitope, this specfic peptide epitope and the monoclonal antibody with specific reactivity against the p24 protein and p24(202-221) can be used in peptide vaccine development and p24 antigen detection from HIV patients.

• 대한바이러스학회지
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• 제26권2호
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• pp.259-267
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• 1996

In order to evaluate the effect of viral load on the prognosis of human immunodeficiency virus-1 (HIV-1)-infected individuals, immune complex dissociated (ICD) serum p24 antigen (p24) by acid treatment was retrospectively measured for 50 HIV-infected patients for 60 months. Among them, 27 patients were p24 positive (p24+) above 25pg/ml for $40.4{\pm}12$ months and 23 patients were negative (p24-). Follow-up periods from HIV diagnosis were $63.0{\pm}19$ months (range; 40-112) for the p24+ and $68.4{\pm}19$ months (range; 38-106) for the p24-, respectively (P>0.05)Mean CD4+T cell counts in the p24+ group decreased from $473{\pm}$277/ul (median;373) to $157{\pm}150/ul$ (median; 111) for $60{\pm}16$ months (5.3/month P280/ul (median; 476) to $432{\pm}285/ul$ (median;382) for $63{\pm}19$ months (2.5/month, P<0.01). From CD4+T cell count >200/ul, the patient who progressed to AIDS of <200/ul were 13 of 23 (56%) in the p24+ and 4 of 22 (18%) in p24-, respectively (p<0.01). And the number of death in two groups were 6 (22%) and 1 (4%), respectively (p<0.01). Presumed survival in two groups were about 12 and 24.5 years. These data suggest that viral load itself be very important for the prognosis of HIV-infected patients.

• BMB Reports
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• 제31권5호
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• pp.432-443
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• 1998

The poliovirus Sabin 1 strain has features that make it a particularly attractive live recombinant mucosal vaccine vehicle. Sabin 1 cDNA was manipulated to have multiple cloning sites and a viral specific 3C-protease cutting site at the N-terminal end of the polyprotein. The gene for the N-terminal 169 amino acids of the HIV-1 p24 was cloned into the multiple cloning site of the manipulated Sabin cDNA. A recombinant progeny virus was produced from HeLa cells when it was transfected with the RNA synthesized from the p24-Sabin chimeric cDNA. The recombinant progeny virus expresses substantial amounts of the HIV-1 p24 protein, which was clearly detected in the infected cell lysates and culture supernatants in Western blot experiments with rabbit anti-p24 serum and AIDS patients' sera. Differing from the Mahoney strain, the recombinant Sabin 1 poliovirus maintained the foreign gene stably during the subsequent passages. Replication capacity was about 1 to 1.5 log lower than that of the wild-type Sabin 1. Other physicochemical stability characteristics of the recombinant virus were similar to that of the wild-type Sabin 1. These results suggest that the manipulated Sabin 1 poliovirus can be used as a live viral vaccine vector for the development of mucosal vaccines.

• 대한미생물학회지
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• 제35권1호
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• pp.9-18
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• 2000

Immunological mechanisms involving the release of inflammatory factors by HIV-1 infected microglia in the brain have been implicated in the pathogenesis of HIV dementia (HIVD). Since the regulation of matrix metalloproteinases (MMPs) activity can be influenced by variety of inflammatory mediators, this study was undertaken to look for a correlation between the MMP-9 release and the production of TNF-${\alpha}$ in response to HIV-1 p24 in the human monocyte cell line THP-1 as a model for microglia. First, it was shown that HIV-l core p24 antigen induced THP-1 to secrete MMP-9 in a dose response manner while it elicited a little effect on MMP-2 release in human astroglial cell line T98G. Next, it was found that p24 induced THP-1 to secrete TNF-${\alpha}$ without prior differentiation into macrophages by phorbol myristate acetate (PMA) treatment. Furthermore, anti-TNF-${\alpha}$ neutralizing antibodies significantly blocked p24-induced MMP-9 release in a dose dependent manner. Our data indicate that p24 antigen induces monocytic MMP-9 release by triggering up-regulation of TNF-${\alpha}$ secretion.

• 대한바이러스학회지
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• 제27권1호
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• pp.77-86
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• 1997

To examine AZT resistance of HIV-1 isolates from AZT treated or untreated Korean, several biological characteristics such as syncytium formation, HIV-1 reverse transcriptase activity and the p24 antigen production in MT-2 cells infected with 4 HIV-1 isolates were determined. As controls, we tested HIV-1 HTLV-IIIB and pre-drug isolate as AZT susceptible strains, in addition to HIV-1 RTMC/MT-2 and post-drug isolate as AZT resistant strains. When the inoculum size of HIV-1 was 300 $TCID_{50}$/well and 100 $TCID_{50}$/well, the AZT susceptibility of AZT untreated HIV-1 isolates 8806 and 9571 were similar to that of HIV-1 HTLV-IIIB and AZT-susceptible HIV-1 strains. When we evaluated AZT resistance of isolates HIV-1 8812 and 9113 treated with AZT for 36 months by observation of syncytium formation, HIV-1 8812 showed resistance simillar to that of HIV-1 RTMC/MT-2 strain forming syncytium up to AZT $1{\mu}g/ml$, and HIV-1 9113 showed resistance identical with that of AZT-resistant HIV-1 strain which formed syncytium up to AZT $10\;{\mu}g/ml$. Especially, when we evaluated AZT resistance by HIV-1 reverse transcriptase activity and the p24 antigen production, HIV-1 isolates 8812 and 9113 showed much higher resistance (>10 - 200 fold) compared with HIV-1 RTMC/MT-2 and AZT-resistant HIV-1 strain.

• 대한바이러스학회지
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• 제28권1호
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• pp.21-30
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• 1998

Synthetic genes encoding the gag p24 and the part of the envelope protein gp41 of the human immunodeficiency virus (HIV-1) were cloned and overexpressed as fusion proteins in Escherichia coli, using an expression vector carrying T7 promoter and the poly-histidine leader, sequence. The overexpressed p24 fusion protein was purified by centrifugation, Ni-affinity chromatography and CM-sepharose chromatography. The overexpressed gp41 fusion protein was purified by centrifugation, $C_4$ chromatography and DEAE-sepharose chromatography. The purified fusion proteins showed a high level of purity and immunoreactivity in SDS-polyacrylamide gel electrophoresis and western blot analysis. These results suggest that this prokaryotic - expression purification method is suitable for obtaining a large amount of the viral antigen which may be useful for screening of antibodies to HIV-1 in human blood samples.

• 미생물학회지
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• 제29권1호
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• pp.1-7
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• 1991

Human immunodeficiency virus type 1 (HIV-1) causes a deadly infectious disease, Acquired Immunodeficiency Syndrome (ADIS). As a first step to develop a reliable and fast diagnostic procedure for HIV-1 infection, we cloned various immunodominant epitopes of HIV-1 in bacterial expression vectors containing tac or trp promoter. While the protein level of direct expression of gp160 was low, trp E fused gp120, gp41 and p17-p24 were produced at high levels (15-30% of total bacterial proteins) in E. coli. Since gp120 and gp41 contain relatively conserved regions which can react with antibodies in the plasma from most of HIV-1 infected individuals, these expression clones were used for large preparations of HIV-1 antigens.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1993년도 제2회 신약개발 연구발표회 초록집
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• pp.80-80
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• 1993

(목적) 본 연구에서 사용된 바이러스는 HIV-1 nef유전자가 일부 삭제되고 대신 Chloramphenicol acetyltransferase(CAT)가 pSVCAT recombinant 바이러스다. 이러한 recombinant 바이러스를 사용하는 이유는 첫째, CAT activity가 매우 민감하므로 바이러스의 복제억제 정도를 정확하게 측정 할 수 있고 둘째, simian immunodeficiency virus(SIV)의 경우 nef 유전가 in vivo에서는 바이러스의 복제에 필수적이므로 HIV가 SIV와 유사한 것으로 미루어 본 연구에서 사용되는 recombinant SVCAT 바이러스가 안전한 것으로 고려되기 때문이다. (방법) 특히 화합물이 HIV-1의 복제에 얼마나 영향이 있는가는 1) 어느정 도의 virus inoculm을 넣었는지 2) 사용하는 cell line 3) 사용한 cell line의 infection kinetics 4) 실험의 지속기간 5) 테스트하는 assay의 sensitivity에 의존한다. 따라서 $10^{5}$ cell의 H9과 sup T1을 24 well plate에 넣고 sup T1 cell line의 경우 3일 후 항 화합물에 의한 syncytia 형성 및 CAT activity의 억제정도를 현재 AIDS drug으로 쓰이고 있는 Zidovudine을 control로 비교 관찰하였다. H9 cell line의 경우 3일 간격으로 media의 3/4을 fresh media로 바꾸어 주고 9일 후 CAT assay를 하였다. 이러한 assay에서 activity를 보이는 화합물을 reverse transcriptase와 P24 ELISA assay를 재확인하였다.다.

• Journal of Microbiology and Biotechnology
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• 제10권5호
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• pp.663-669
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• 2000

The fur (ferric uptake regulation) expression vector pMON2064 was modified to produce a Fur-fusion expression vector. A kinker site, factor Xa cleavage site, and several restriction endonuclease sites were introduced to facilitate easy cloning and isolating of the fusion protein. The resulting fusion expression vector, pMONSTER, was then used to make fusion expression vector, pMONSTER, was then used to make fusion proteins with $\beta$-galactosidase and the protease of the human immunodeficiency virus type 1 (HIV-1 PR). Strain SW4020 harboring the Fur $\beta$-galactosidase fusion vector produced blue colonies on a 5-bromo-4-chloro-3-indolyl-$\beta$-D-galactoside plate and the resulting 133 kDa fusion protein reacted with an anti-Fur antibody. The strain harboring the Fur-HIV-1 PR fusion vector produced a 29 kDa fusion protein, which also reacted with an anti-Fur antibody. The Fur-HIV-1 PR fusion protein was purified by a single column application that was designed to isolate the Fur protein. The purified Fur-HIV-1 PR fusion protein digested with factor Xa cleaved a recombinant Gag protein to release smaller fragments, including a p24 capsid protein. The Fur-HIV-1 PR fusion protein itself did not exhibit any proteolytic activity.

• 대한바이러스학회지
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• 제28권4호
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• pp.347-358
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• 1998

To analyze the correlation between biological phenotypes of HIV-1 isolates and disease progression, we selected 9 long-term non-progressors (LTNP) and 12 rapid progressors (RP) from HIV-1 infected Korean. We isolated HIV-1 isolates by culture of PBMC of LTNP and RP with normal PBMC and measured HIV-1 p24 antigen production. The HIV-1 isolation rate from LTNP was 55.6% (5/9). And 4 HIV-1 LTNP isolates were non-syncytium inducing (NSI) phenotype and showed slow/low replication. The HIV-1 isolation rate from RP was 91.7% (11/12) which was higher than that from LTNP. Besides 3 RP HIV-1 isolates which showed syncytium inducing (SI) phenotype, 8 RP HIV-1 isolates showed NSI phenotype in normal PBMC and MT-2 cell line. All RP HIV-1 isolates replicated more rapidly than LTNP HIV-1 isolates. Comparing the replication kinetics and syncytium forming capacity of HIV-1 isolates from LTNP and RP, we suggest that the difference of biological phenotype of HIV-1 isolates could be related with disease progression of HIV-1 infected persons.