• 대한약학회:학술대회논문집
• /
• 대한약학회 2001년도 Proceedings of the Pharmaceutical Society of Korea
• /
• pp.169.1-169.1
• /
• 2001

• Journal of Microbiology and Biotechnology
• /
• 제21권5호
• /
• pp.529-535
• /
• 2011

Single-chain variable fragment (scFv) is a fusion protein of the variable regions of the heavy (VH) and light (VL) chains of immunoglobulin, connected with a short linker peptide of 10 to about 20 amino acids. In this study, the scFv of a monoclonal antibody against the third domain of human CD4 was cloned from OKT4 hybridoma cells using the phage display technique and produced in E. coli. The expression, production, and purification of anti-CD4 scFv were tested using SDS-PAGE and Western blot, and the specificity of anti-CD4 scFv was examined using ELISA. A 31 kDa recombinant anti-CD4 scFv was expressed and produced in bacteria, which was confirmed by SDS-PAGE and Western blot assays. Sequence analysis proved the ScFv structure of the construct. It was able to bind to CD4 in quality ELISA assay. The canonical structure of anti-CD4 scFv antibody was obtained using the SWISS_MODEL bioinformatics tool for comparing with the scFv general structure. To the best of our knowledge, this is the first report for generating scFv against human CD4 antigen. Engineered anti-CD4 scFv could be used in immunological studies, including fluorochrome conjugation, bispecific antibody production, bifunctional protein synthesis, and other genetic engineering manipulations. Since the binding site of our product is domain 3 (D3) of the CD4 molecule and different from the CD4 immunological main domain, including D1 and D2, further studies are needed to evaluate the anti-CD4 scFv potential for diagnostic and therapeutic applications.

• 약학회지
• /
• 제57권2호
• /
• pp.125-131
• /
• 2013

Biologically active peptides, including growth factors and cytokines, participate in various biological processes in human skin. They could provide a great advantage of maintaining healthy skin. Many peptide growth factors like epidermal growth factor (EGF) and human growth hormone (hGH) have been used in cosmetic formulations. The delivery of peptide growth factors across the Stratum corneum, however, seems not sufficient because of their physical properties such as high molecular weight and hydrophilicity. So increasing the penetration of growth factors of interest into skin would be a major concern for ensuring their maximum biological efficacy. In this study, we have identified several skin penetration-enhancing peptides which facilitate delivery of growth factors, when fused at N-terminus of the target protein, into skin. For efficient and rapid screening, we constructed a skin-penetrating assay system using Franz cell and porcine skin. Next, we carried out phage display screening using M-13 bacteriophage with random 12 -amino acid library on its coat protein P3 on that system. After several selection rounds, peptide sequences facilitate the penetration of phages through the porcine skin were identified from a large population of phages. We found that phages with the most potent peptide (S3-2, NGSLNTHLAPIL) could penetrate the porcine skin eight times more than those with control peptide (12 mino acids scrambled peptide). Furthermore, growth factors conjugated with S3-2 peptide penetrate porcine skin three to five times efficiently than non-conjugated growth factors. In conclusion, our data shows that the skin penetration-enhancing peptide we have characterized could increase the delivery of growth factors and is useful for cosmeceutical application.

• 대한금속재료학회지
• /
• 제46권3호
• /
• pp.131-134
• /
• 2008

The magnetism of$Fe_3O_4$ nanoparticles was applied to magnetic bio-panning process for finding specific sequences against $Fe_3O_4$ crystal phase. Vibrating sample magnetometer (VSM) measurement showed that the coercivity of 30 Oe and the saturation magnetization of 55 emu/g were sufficient in controlling particle movement and magnetizing particles in the media, respectively. This ferrimagnetism of nanoparticles practically enhanced panning efficiency by exaggerating centrifuge step and preventing particle loss. Sequencing results showed that histidine which was commonly found in peptide sequences played an important role in the binding onto $Fe_3O_4$ nanoparticle surface. However, various possible motifs were also observed from several neighboring amino acids of histidine.

• Journal of Microbiology and Biotechnology
• /
• 제16권11호
• /
• pp.1784-1790
• /
• 2006

Bacillus anthracis is a causative agent of anthrax. Anthrax toxins are composed of a protective antigen (PA), lethal factor (LF), and edema factor (EF), in which the PA is a central mediator for the delivery of the two enzymatic moieties LF and EF. Therefore, the PA has been an attractive target in the prevention and vaccinization for anthrax toxin. Recently, it has been reported that the molecule consisting of multiple copies of PA-binding peptide, covalently linked to a flexible polymer backbone, blocked intoxification of anthrax toxin in an animal model. In the present study, we have screened novel diverse peptides that bind to PA with a high affinity (picomolar range) from an M13 peptide display library and characterized the binding regions of the peptides. Our works provide a basis to develop novel potent inhibitors or diagnostic probes with a diverse polyvalence.

• 한국미생물·생명공학회지
• /
• 제35권4호
• /
• pp.278-284
• /
• 2007

생체 염 농도에서도 항균활성을 유지할 수 있는 선형 ${\alpha}$-helical 항균 펩타이드를 M13 펩타이드 라이브러리로부터 탐색할 수 있는 새로운 방법을 개발하였다. M13의 pIII은 magainin 유도체인 MSI-344와 indolicidin과 융합된 상태에서도 파아지의 viability에 영향을 주지 않는 것으로 보아, MSI-344와 indolicidin의 대장균에 대한 독성을 중화할 수 있는 것으로 판단되며, 따라서 대장균에서 항균 펩타이드 라이브러리의 제조가 가능함을 증명하였다. 선형 항균 펩타이드의 보존된 부위를 바탕으로, 13개의 아미노산 잔기로 구성된 semi-combinatorial 항균 펩타이드 라이브러리를 M13를 이용하여 제조하였다. 제조된 파아지 라이브러리는 먼저 적혈구에 흡착시켜, 높은 용혈 역가를 가질 가능성이 있는 파아지를 제거한 후, 높은 염 농도에서 Pseudomonas aeruginosa와 Staphylococcus aureus에 흡착할 수 있는 파아지를 탐색하였다. 탐색된 펩타이드들은 염이 없는 조건에서는 비교적 낮은 항균 역가를 보였지만, P06와 S18 펩타이드의 경우, 생체 염 농도보다 높은 150 mM $Na^+$, 2 mM $Mg^{2+}$, 2 mM $Ca^{2+}$의 조건에서도 항균 역가가 영향을 받지 않았으며, 심각한 용혈 역가 또한 보이지 않았다. 본 연구에서 개발한 대상 세균에 대한 흡착능력을 이용한 탐색방법은 salt-tolerant antimicrobial peptide의 개발의 새로운 가능성을 제시하였다.

• 한국생물공학회:학술대회논문집
• /
• 한국생물공학회 2005년도 생물공학의 동향(XVI)
• /
• pp.393-393
• /
• 2005

• 한국생물공학회:학술대회논문집
• /
• 한국생물공학회 2003년도 생물공학의 동향(XII)
• /
• pp.749-749
• /
• 2003

• 한국생물물리학회:학술대회논문집
• /
• 한국생물물리학회 1998년도 학술발표회
• /
• pp.11-11
• /
• 1998

Zinc fingers of Cys$_2$His$_2$ class constitute one of the most common DNA binding motifs in eukaryotes. Unlike other DNA binding motifs, zinc finger proteins recognize very diverse DNA sites, and their sequence specificities can be systematically changed by phage display.(omitted)

• Journal of Microbiology and Biotechnology
• /
• 제24권5호
• /
• pp.704-713
• /
• 2014

The highly pathogenic avian influenza A (HPAI) viruses of the H5N1 subtype infect poultry and have also been spreading to humans. Although new antiviral drugs and vaccinations can be effective, rapid detection would be more efficient to control the outbreak of infections. In this study, a phage-display library was applied to select antibody fragments for HPAI strain A/Hubei/1/2010. As a result, three clones were selected and sequenced. A hemagglutinin inhibition assay of the three scFvs revealed that none exhibited hemagglutination inhibition activity towards the H5N1 virus, yet they showed a higher binding affinity for several HPAI H5N1 strains compared with other influenza viruses. An ELISA confirmed that the HA protein was the target of the scFvs, and the results of a protein structure simulation showed that all the selected scFvs bound to the HA2 subunit of the HA protein. In conclusion, the three selected scFVs could be useful for developing a specific detection tool for the surveillance of HPAI epidemic strains.