Refolding of autodisplayed anti-NEF scFv through oxidation with glutathione for immunosensors

Bong, Ji-Hong,Song, Hyun-Woo,Kim, Tae-Hun,Kang, Min-Jung,Jose, Joachim,Pyun, Jae-Chul Elsevier Applied Science 2018 Biosensors & bioelectronics Vol. No.

<P><B>Abstract</B></P> <P>In this study, a single-domain antibody against negative regulatory factor (anti-NEF scFv) was autodisplayed on the outer membrane of <I>Escherichia coli</I> and used to detect NEF in an immunoassay based on fluorescence-activated cell sorting, enzyme-linked immunosorbent assay, and surface plasmon resonance biosensors. Next, the autodisplayed single-domain antibody was oxidized to form disulfide bonds by using glutathione, and the change in NEF-binding activity of anti-NEF scFv was analyzed by fluorescence-activated cell sorting-based immunoassay, chromogenic immunoassay, and surface plasmon resonance biosensor. For each type of immunoassays the anti-NEF scFv on the isolated outer membrane showed more NEF binding activity after the disulfide bond formation by glutathione. To determine the role of cysteines in anti-NEF scFv, three mutants were prepared, and the NEF binding activity of mutants was compared with that of wild-type anti-NEF scFv in a competitive immunoassay based on FACS. In these mutant studies, the refolding process of autodisplayed anti-NEF scFv by following oxidation via GSH/GSSG revealed that disulfide bonds formed and increased NEF binding activity.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Anti-NEF scFv was autodisplayed on the outer membrane (OM) of <I>Escherichia coli</I>. </LI> <LI> Autodisplayed anti-NEF scFv was oxidized to form disulfide bonds by using glutathione. </LI> <LI> The activity of scFv was analyzed by FACS-based immunoassay and SPR biosensor. </LI> <LI> Mutants were made to confirm the formation of disulfides after the reaction of glutation. </LI> </UL> </P>

NeH_2, NeF_2, NeCl_2, NeCIF에 대한 이론 연구

손양숙,성은모 충북대학교 과학교육연구소 2001 과학교육연구논총 Vol.17 No.1

The ab initio molecular orbital calculation of NeH_2, NeF_2, NeCl_2 and NeC1F are carried out by Moller-Plesset method with 6-31G* basis set, and the structures of the minimum energy points were obtained. Each of these van der Waals complexes showed the linear form as the most stable structure, and T-shaped complexes are also stable for NeCl_2 and NeCIF. The binding energies were calculated for all of those van der Waals complexes, and showed the reasonable agreements with the experimental values.

Bayesian Estimation of the Difference between Two Means of Natural Exponential Families with Quadratic Functions

Cha,Myung-Soo 慶星大學校 1991 論文集 Vol.12 No.2

The problem of allocationg a fixed number of trials sequentially between tow independent populations within al NEF-QVF so as to estimate the difference between their means is considered. Assuming conjugate prior on the parameters a heuristic allocation rule applied and its asymptotic optimality is shown.

T 세포 특이적 전사인자인 LyF-1과 HIV-1 Nef의 상호 작용

이미선(Mi Seon Lee),이경화(Kyoung Hoa Lee),김정우(Jung Woo Kim) 대한바이러스학회 2000 Journal of Bacteriology and Virology Vol.30 No.3

Korean red ginseng attenuates HIV-1 in vivo; High frequency of grossly deleted nef genes in HIV-1 infected long-term slow progressors treated with Korean red ginseng

Young K. Cho(조영걸),Ji Y. Lim,You S. Jung,Sun K. Oh,Hee J. Lee,Heungsup Sung 고려인삼학회 2006 고려인삼학회 학술대회 Vol.- No.-

To investigate the association between Korean red ginseng (KRG) intake in HIV-1 infected patients and occurrence of grossly deleted nef genes (g?nef), we characterized nef genes in 10 long-term slow progressors (LTSP) infected with HIV-1 subtype B and 34 control patients. LTSP was defined whose the annual decrease in CD4 T cells was less than 20/μl over 10 years in the absence of antiretroviral therapy. They were treated with KRG for a prolonged period. Nef genes were amplified from peripheral blood mononuclear cells (PBMC) using nested PCR and products were sequenced directly. Patient CD4 T cell counts decreased from 444 ± 207/㎕ to 294 ± 177/㎕ over 136 ± 23 months of KRG intake. This corresponds to an annual decrease in the level of CD4 T cells of 13.3/㎕. A total of 479 nef genes were amplified from 137 PBMC samples. Nine out of the 10 patients, 47 (34.3%) out of the 137 samples, and 92 out of the 479 genes revealed g?nef. The deletion extended outside the nef gene in 25 g?nef obtained from 6 patients. The proportion of samples with g?nef (34.3%) was significantly higher than 4.8% in control patients (P < 0.001). In addition, it significantly increased as the duration of KRG intake prolongs (P < 0.01). These data suggest the possibility that occurrence of g?nef might be associated with long?term intake of KRG.

Genetic defects in the nef gene are associated with Korean Red Ginseng intake: monitoring of nef sequence polymorphisms over 20 years

Cho, Young-Keol,Kim, Jung-Eun,Woo, Jun-Hee The Korean Society of Ginseng 2017 Journal of Ginseng Research Vol.41 No.2

Background: The presence of gross deletions in the human immunodeficiency virus nef gene ($g{\Delta}nef$) is associated with long-term nonprogression of infected patients. Here, we investigated how quickly genetic defects in the nef gene are associated with Korean Red Ginseng (KRG) intakein 10 long-term slow progressors. Methods: This study was divided into three phases over a 20-yr period; baseline, KRG intake alone, and KRG plus highly active antiretroviral therapy (ART). nef gene amplicons were obtained using reverse transcription polymerase chain reaction (PCR) and nested PCR from 10 long-term slow progressors (n = 1,396), and nested PCR from 36 control patients (n = 198), and 28 ART patients (n = 157), and these were then sequenced. The proportion of $g{\Delta}nef$, premature stop codons, and not in-frame insertion or deletion of a nucleotide was compared between three phases, control, and ART patients. Results: The proportion of defective nef genes was significantly higher in on-KRG patients (15.6%) than in baseline (5.7%), control (5.6%), on-KRG plus ART phase (7.8%), and on-ART patients (6.6%; p < 0.01). Small in-frame deletions or insertions were significantly more frequent among patients treated with KRG alone compared with controls (p < 0.01). Significantly fewer instances of genetic defects were detected in samples taken during the KRG plus ART phase (7.8%; p < 0.01). The earliest defects detected were $g{\Delta}nef$ and small in-frame deletions after 7 mo and 67 mo of KRG intake, respectively. Conclusion: KRG treatment might induce genetic defects in the nef gene. This report provides new insight into the importance of genetic defects in the pathogenesis of AIDS.

Dosage and Duration Effects of Korean Red Ginseng Intake on Frequency of Gross Deletions in the nef Gene

Cho, Young-Keol,Jung, You-Sun The Korean Society of Ginseng 2010 Journal of Ginseng Research Vol.34 No.3

In the present study, we investigated whether a gross deletion in the nef gene ($g{\Delta}nef$) is induced by Korean red ginseng (KRG) intake. Ten patients were treated with KRG powder for 3 years in the absence of antiretroviral drug therapy. On average, $3,555{\pm}1,042\;g$ KRG was administered per person over $36.1{\pm}2.4$ months. There was a mild decrease in CD4 T cell count ($75{\pm}110/{\mu}L$) over the $36.1{\pm}2.4$ months (p = 0.059). We obtained 355 nef amplicons using 71 peripheral blood mononuclear cell samples over a 3-year period. All ten patients exhibited g${\Delta}$nef (range, 3.2 to 45.9%). At baseline, 3 of 78 amplicons (3.8%) exhibited $g{\Delta}nef$, whereas 18.8% (52/277) revealed $g{\Delta}nef$ during KRG-intake (p<0.001). The proportion of $g{\Delta}nef$ was significantly correlated with monthly dose of KRG (r=0.89, p<0.001). The median time for first detection of $g{\Delta}nef$ was 13 months. In conclusion, our data show that $g{\Delta}nef$ is inducible by KRG intake and its proportion is dependent on the duration of KRG intake and dose of KRG.

국내 감염성 질환 병원체의 성상 분석을 통한 표준 실험실 체계 구축 : 우리나라의 AIDS reagent 프로그램 개발 및 국내 HIV/AIDS 물질 자원화

강춘,남정구,김성순,최병선,김옥진,김재인,이성래,구본기,서순덕,이주실,조해월 국립보건연구원 1999 국립보건원보 Vol.36 No.-

Dosage and Duration Effects of Korean Red Ginseng Intake on Frequency of Gross Deletions in the nef Gene

Young Keol Cho,You Sun Jung 고려인삼학회 2010 Journal of Ginseng Research Vol.34 No.3

In the present study, we investigated whether a gross deletion in the nef gene (g?nef) is induced by Korean red ginseng (KRG) intake. Ten patients were treated with KRG powder for 3 years in the absence of antiretroviral drug therapy. On average, 3,555 ± 1,042 g KRG was administered per person over 36.1±2.4 months. There was a mild decrease in CD4 T cell count (75 ± 110/mL) over the 36.1±2.4 months (p = 0.059). We obtained 355 nef amplicons using 71 peripheral blood mononuclear cell samples over a 3?year period. All ten patients exhibited g?nef (range, 3.2 to 45.9%). At baseline, 3 of 78 amplicons (3.8%) exhibited g?nef, whereas 18.8% (52/277) revealed g?nef during KRG?intake (p < 0.001). The proportion of g?nef was significantly correlated with monthly dose of KRG (r = 0.89, p < 0.001). The median time for first detection of g?nef was 13 months. In conclusion, our data show that g?nef is inducible by KRG intake and its proportion is dependent on the duration of KRG intake and dose of KRG.

Genetic defects in the nef gene are associated with Korean Red Ginseng intake

Young-Keol Cho,Jung-Eun Kim,Jun-Hee Woo 고려인삼학회 2017 Journal of Ginseng Research Vol.41 No.2

Background: The presence of gross deletions in the human immunodeficiency virus nef gene (gDnef) is associated with long-term nonprogression of infected patients. Here, we investigated how quickly genetic defects inthe nef gene are associatedwithKoreanRedGinseng (KRG) intake in10 long-termslowprogressors. Methods: This study was divided into three phases over a 20-yr period; baseline, KRG intake alone, and KRG plus highly active antiretroviral therapy (ART). nef gene amplicons were obtained using reverse transcription polymerase chain reaction (PCR) and nested PCR from 10 long-term slow progressors (n ¼ 1,396), and nested PCR from 36 control patients (n ¼ 198), and 28 ART patients (n ¼ 157), and these were then sequenced. The proportion of gDnef, premature stop codons, and not in-frame insertion or deletion of a nucleotide was compared between three phases, control, and ART patients. Results: The proportion of defective nef genes was significantly higher in on-KRG patients (15.6%) than in baseline (5.7%), control (5.6%), on-KRG plus ART phase (7.8%), and on-ART patients (6.6%; p < 0.01). Small in-frame deletions or insertions were significantly more frequent among patients treated with KRG alone compared with controls (p < 0.01). Significantly fewer instances of genetic defects were detected in samples taken during the KRG plus ART phase (7.8%; p < 0.01). The earliest defects detected were gDnef and small in-frame deletions after 7 mo and 67 mo of KRG intake, respectively. Conclusion: KRG treatment might induce genetic defects in the nef gene. This report provides new insight into the importance of genetic defects in the pathogenesis of AIDS.