• Title/Summary/Keyword: Light chain 3

Search Result 296, Processing Time 0.023 seconds

Periplasmic Expression of a Recombinant Antibody (MabB9) in Escherichia coli

  • Chang, Hae-Choon;Kwak, Ju-Won
    • Journal of Microbiology and Biotechnology
    • /
    • v.7 no.5
    • /
    • pp.299-304
    • /
    • 1997
  • Expression in the periplasm of Escherichia coli of cloned heavy and light chain cDNAs for Fab fragment of a murine monoclonal antibody MabB9 (${\gamma}2b$, K), specific for human plasma apolipoprotein B-100 of LDL, was studied. For the purpose, a vector for two-cistronic expression of the heavy chain cDNA, at the 5' terminus, and light chain cDNA, at the 3' terminus, was constructed using the signal sequences, pelB (for heavy chain) and ompA (for light chain) in a pET vector system. The constructed vector was transformed into E. coli BL21(DE3). The expressed heavy chain (25 kDa) and light chain (23 kDa) of the antibody molecule were detected in total cell extracts as well as in the periplasmic extracts of E. coli.

  • PDF

Identification of the Interaction between Rat Translationally Controlled Tumor Protein/IgE-dependent Histamine Releasing Factor and Myosin Light Chain

  • Kim, Min-Jeong;Jung, Jae-Hoon;Choi, Eung-Chil;Park, Hae-Young;Lee, Kyung-Lim
    • BMB Reports
    • /
    • v.34 no.6
    • /
    • pp.526-530
    • /
    • 2001
  • The translationally controlled tumor protein (TCTP), also known as the IgE-dependent histamine releasing factor (HRF), was used in the yeast two-hybrid system to screen the interacting molecules. We obtained the N-terminus truncated rat fast myosin alkai light chain from the rat skeletal muscle cDNA library in the screening. Since either TCTP/HRF or the myosin light chain is known to be associated with histamine secretion from RBL-2H3 cells, we investigated the possible interaction between rat TCTP/HRF and nonmuscle myosin light chain in these cells. We used affinity chromatography and coimmunoprecipitation. Our data suggests that HRF and the myosin light chain interact, which may play an important role in histamine release in RBL-2H3 cells.

  • PDF

In vitro Propagation of Transgenic Ginsengs Introduced with Ferritin Light Heavy Chain Gene through Single Embryo Culture (Ferritin Light Heavy Chain 유전자가 도입된 인삼형질전환체의 단일배발생을 통한 식물체의 기내증식)

  • 윤영상;김종학;김무성;양덕춘
    • Korean Journal of Plant Resources
    • /
    • v.17 no.2
    • /
    • pp.161-168
    • /
    • 2004
  • Optimal regeneration conditions of ginseng transformants were studied. It has been known that Ferritin Light Heavy Chain (FLHC) gene remove the several heavy metal by combination, store and transport. To obtain the ginseng tolerant to heavy metal, binary vector was introduced in Agrobacterium by tri-parental mating and then Agrobacterium tumefaciens MP90/FLHC was selected on the AB media and MS media containing kanamycin. Explants were co-cultured with Agrobacterium tumefaciens MP90/FLHC, which contained NPT II as a selectable marker, tadpole ferritin heavy chain (FLHC) gene and human ferritin light chain gene and then a number of embryos were induced. The induced embryo transferred to shooting media consisting of MS medium supplemented with GA 10 mg/L. As a result of examination that induced the normal growth of transfomants, transformants showed the equivalent growth in both root and shoot on the media containing the 1/3 MS.

Optical Anisotropy of Polyimide and Polymethacrylate Containing Photocrosslinkable Chalcone Group in the Side Chain under Irradiation of a Linearly Polarized UV Light

  • Choi, Dong-Hoon;Cha, Young-Kwan
    • Bulletin of the Korean Chemical Society
    • /
    • v.23 no.3
    • /
    • pp.469-476
    • /
    • 2002
  • Photocrosslinkable soluble polyimide and polymethacrylate compound were synthesized for studying the optically induced anisotropy of the thin films. Chalcone group was introduced into the side chain unit of two polymers. We observed a photodimerization behavior between the double bonds in the chalcone group and an optical anisotropy of these materials by irradiation of a linearly polarized UV light (LPL). Optical anisotropy of the thin film was also investigated by using polarized UV absorption spectroscopy. The dynamic property of optical anisotropy in photoreactive polyimide was compared to that in polymethacrylate containing chalcone group in the side chain.

Disulfide Bond Bridged Divalent Antibody-Toxin, $(Fab-PE38fl)_2$ with the Toxin PE38 Fused to the Light Chain

  • Won, Jae-Seon;Choe, Mu-Hyeon
    • Journal of Microbiology and Biotechnology
    • /
    • v.18 no.8
    • /
    • pp.1475-1481
    • /
    • 2008
  • B3 antibody specifically binds the $Lewis^Y$-related carbohydrate antigen of many carcinomas, and it is used as a model antibody in this study. In a previous study, the Fab fragment of the antibody was fused to a 38 kDa truncated form of Pseudomonas exotoxin A, PE38, to make Fab-PE38, where PE38 is fused to the Fd fragment of the Fab domain. This parent monomer molecule, Fab-PE38, had no cysteine in the hinge region, and it could not make a disulfide bond to form a disulfide bond bridged homodimer. In this study, we constructed three different kinds of divalent Fab-toxin fusion homodimers where the toxin is fused to the light chain of Fab, $(Fab-PE38fl)_2$. In addition to the PE38 toxin fused to the light chain, these three molecules have different hinge sequences hi, h2, and h3 making Fabh1-, Fabh2-, and Fabh3-PE38fl monomers, respectively. These hinges contain only one cysteine on different positions of the hinge sequence. The disulfide bond between the hinge region of two monomers forms homodimers $(Fabh1-PE38fl)_2$, $(Fabh2-PE38fl)_2$, and $(Fabh3-PE38fl)_2$. The refolding yields of these dimers were 5-16-fold higher than a previously constructed dimer where the PE38 was fused to the Fd fragment $(Fabh2-PE38)_2$ [8]. Our data suggest that the steric repulsion between the two PE38s in $(Fabh1-PE38)_2$ during disulfide bridge formation is relieved by fusing it at the end of the light chain. The best cytotoxicity value of these dimers showed about 2.5-fold higher on an MCF7 cell line than that of the monovalent reference molecule in ng/ml scale, which is 15-fold higher in pM scale.

Relationships between Myosin Light Chain Isoforms, Muscle Fiber Characteristics, and Meat Quality Traits in Porcine Longissimus Muscle

  • Choi, Young-Min;Ryu, Youn-Chul;Lee, Sang-Hoon;Kim, Byoung-Chul
    • Food Science and Biotechnology
    • /
    • v.14 no.5
    • /
    • pp.639-644
    • /
    • 2005
  • The aim of this study was to investigate the effect of the myosin light chain (MLC) isoforms on the muscle fiber characteristics and meat quality traits in porcine longissimus muscle. Pale, soft, exudative (PSE) samples had a lower content of essential light chain (ELC) 1S isoforms and a higher proportion of the fiber type IIB than the reddish-pink, firm, non-exudative (RFN) samples. These compositions suggest that the PSE pork has a higher glycolytic and a lower oxidative capacity than the RFN pork. Therefore, these characteristics of PSE pork might affect the metabolic rate and meat quality traits, including protein solubility. In addition, the indicator traits of the postmortem metabolic rate were related to the ELC 1F/3F ratio ($pH_{45\;min}$: r = -0.43, P < 0.001; R-value: r = 0.53, P < 0.001). These results suggest that the MLC isoform composition can affect the postmortem metabolic rate and meat quality traits.

Production, Characterization, and Variable Region Analysis of Monoclonal Antibodies Specific for Hepatitis B Virus S Antigen (Hepatitis B Virus의 S항원에 특이적인 단세포군 항체 생산, 특성 연구 및 가변지역유전자 분석)

  • Song, Moo-Young;Kim, Chang-Seok;Park, Sang-Koo;Lee, Jae-Sun;Yoo, Tae-Hyoung;Ko, In-Young
    • IMMUNE NETWORK
    • /
    • v.3 no.4
    • /
    • pp.281-286
    • /
    • 2003
  • Background: Hepatitis B virus (HBV) infection is one of the worldwide public health problem affecting about 300 million people. The envelope protein of HBV consists of three components known as preS1, preS2, and S antigen. According to the recent study, anti-HBs Ab showed effective neutralization ability against HBV from chronic hepatitis B and liver transplant patients, suggesting the possible development of therapeutic antibody. Methods: Spleen cells immunized with S antigen of HBV were fused with myeloma cell line to obtain HBsAg specific monoclonal antibodies. High affinity antibodies against HBsAg (adr, ad and ay type) were selected by competitive ELISA method. Nucleotide sequence of the variable regions of monoclonal antibodies was analyzed by RT-PCR followed by conventional sequencing method. Results: We produced 14 murine monoclonal antibodies which recognize S antigen of HBV. Two of them, A9-11 and C6-9 showed the highest affinity. The sequence analysis of A9-11 revealed that variable regions of the heavy chain and light chains are members of mouse heavy chain I (B) and light chain lambda 1, respectively. Likewise, the sequence analysis of C6-9 revealed that variable regions of the heavy chain and light chains are members of mouse heavy chain II (B) and light chain kappa 1, respectively. Neutralization assay showed that A9-11 and C6-9 effectively neutralize the HBV infection. Conclusion: These results suggest that A9-11 and C6-9 mouse monoclonal antibodies can be used for the development of therapeutic antibody for HBV infection.

Construction and Characterization of a Single-Chain Immunoglobulin

  • Kim, Youn-Kyu;Choi, In-Hak;Ryu, Chun-Jeih;Hong, Hyo-Jeong
    • BMB Reports
    • /
    • v.30 no.3
    • /
    • pp.177-181
    • /
    • 1997
  • We constructed a single-chain immunoglobulin in which the carboxyl end of the heavy chain variable domain is covalently joined to the amino terminus of the light chain variable domain via peptide linker and the carboxyl end of the light chain variable domain is linked to human ${\gamma}1$ Fc region through the hinge region. The molecule was expressed in Chinese hamster ovary cells, assembled into a dimeric molecule and secreted into the culture medium. The dimeric molecule (2E11) was purified from the culture supernatant by affinity chromatography on Protein G-Sepharose column. The size of the unreduced or reduced protein was the expected molecular weight of approximately 120 or 60 kDa, respectively, as assessed by SDS-polyacrylamide gel electrophoresis. The antigen-binding affinity of 2E11 was almost the same as that of a native antibody counterpart (CS131A), suggesting that the single-chain immunoglobulin may function like a native antibody.

  • PDF

Analysis of Immunoglobulin λ Light Chain Repertoire in Systemic Lupus Erythematosus (루푸스 환자의 면역글로불린 λ 경쇄 레파토리 분석)

  • Chang, Ji Eun;Lee, Jisoo
    • IMMUNE NETWORK
    • /
    • v.3 no.3
    • /
    • pp.227-234
    • /
    • 2003
  • Background: Immunoglobulin (Ig) light chain repertoire has been implicated as a critical determinant in regulation of autoreactive B cells and production of pathogenic anti-DNA antibodies in systemic lupus erythematosus (SLE). We analyzed the impact of Ig ${\lambda}$ chain repertoire on development of autoimmunity in patients with SLE. Methods: We obtained genomic DNA from individual peripheral CD19+ B cells of 3 untreated active SLE patients, and amplified $V{\lambda}$ rearrangements from each single cell by polymerase chain reaction. Results: A total number of 208 $V{\lambda}J{\lambda}$ rearrangements were analyzed. Analyzed sequences included 158 productive rearrangements and 50 nonproductive rearrangements. The differences in $V{\lambda}$ gene usage in the productive and nonproductive repertoire of SLE patients were found compared to the non-autoimmune individuals. $V{\lambda}$ gene, 9A was significantly overrepresented in nonproducative repertoire of SLE patients (P=0.016). In the productive repertoire, $V{\lambda}$ genes, 3L and 1E were found more often in the SLE patients (P=0.001, P=0.043). When the productive and the nonproductive repertoires were compared, 9A was found significantly less in the productive repertoire in the SLE patients (P=0.000). There were no significant differences in the $J{\lambda}$ gene usage between SLE patients and non-autoimmune individuals, but $J{\lambda}2/3$ gene was the most frequently used in SLE, whereas $J{\lambda}7$ gene was the most frequently used in the normal subjects. In the productive SLE $V{\lambda}$ repertoire, 9.4% of the total sequences employed identical CDR3. It was particularly striking to find 7 identical versions of the 1G-$J{\lambda}2/3$ $V{\lambda}J{\lambda}$ rearrangements from one patient and 3 of the same sequence from another patient. Notably, identical $V{\lambda}$ junctions in the SLE patients utilized significantly more homologous joining compared to $V{\lambda}$ junctions of the normal adults (P=0.044). Conclusion: These data demonstrate regulation of ${\lambda}$ light chain expression in the SLE patients by selection of unique $V{\lambda}$ genes. Also, biased selection and clonal expansion of particular $V{\lambda}$ rearrangements are apparent in the SLE ${\lambda}$ repertoire.

Dynamic Characteristic Evaluation of the Bucket Elevator Chain Pin and Plate (버킷 엘리베이터 체인의 동특성 평가)

  • Kim, Chang Uk;Lee, Dong Woo;Park, Seung Bin;Song, Jung Il
    • Journal of the Korean Society for Precision Engineering
    • /
    • v.34 no.3
    • /
    • pp.211-215
    • /
    • 2017
  • This research analyzes bucket elevator roller chain pins by finite element (FE) analysis and static structural analysis for a lightweight pin design. The stress distribution of light weight roller chain pins under static load is analyzed for safety factors and damping effect. The results show that the stress distribution is higher on the plate than on the bush pin. In order to compare experimental and FE analysis results, a light weight design approach was used to produce a prototype base pin. Because the inner diameter of the pin was different, the impact damping effect was most appropriate when the inner diameter was 34.05 mm, and it is used as basic research data on the impact of the roller chain and sprocket.