• Archives of Pharmacal Research
• /
• v.25 no.6
• /
• pp.801-806
• /
• 2002

Cyclicpeptides are important targets in peptide synthesis because of their interesting biological properties. Constraining highly flexible linear peptides by cyclization is one of the mostly widely used approaches to define the bioactive conformation of peptides. Cyclic peptides often have increased receptor affinity and metabolic stability over their linear counterparts. We carried out virtual screening experiment via docking in order to understand the interaction between HLE-Human Leukocyte Elastase and ligand peptide and to identify the sequence that can be a target in various ligand peptides. We made cyclic peptides as a target base on Metlle-Phe sequence having affinity for ligand and receptor active site docking. There are three ways to cyclize certain sequences of amino acids such as Met-lie-Phe-Gly-Ile. First is head-to-tail cyclization method, linking between N-terminal and C-terminal. Second method utilizes amino acid side chain such as thiol functional group in Cys, making a thioether bond. The last one includes an application of resin-substituted amino acids in solid phase reaction. Among the three methods, solid phase reaction showed the greatest yield. Macrocyclization of Fmoc-Met-Ile-Phe-Gly-Ile-OBn after cleavage of Fmoc protection in solution phase was carried out to give macrocyclic compound 5 in about 7% yield. In the contrast with solution phase reaction, solid phase reaction for macrocyclization of Met-Ile-Phe-Gly-Ile-Asp-Tentagel in normal concentrated condition gave macrocyclic compound 7 in more than 35% yield.

• Journal of the Korean Chemical Society
• /
• v.32 no.5
• /
• pp.483-494
• /
• 1988

The adsorption behavior of aromatic acids on Amberlite XAD-4 resin was investigated by measuring the distribution coefficient by batch method. The adsorption of solutes on XAD-4 was affected by the several factors such as, analyte concentration, the pH of solution and concentration of pairing ion. The enhanced adsorption of solutes on XAD-4 in the presence of tetraalkylammonium salt as an ion pairing reagent, referred to as ion interaction, was suggested to follow a double layer model where the pairing ion occupies a primary layer at the adsorbent while the solute anion and other anions in the system comlpete for the secondary layer. Therefore, the ability of an ion pairing reagent to enhance solute adsorption depended significantly on the type and concentration of counter-ion and co-anion accompanying the ion pairing reagent or salt used for ionic strength control. In addition, a good linear relationship between the logarithm of capacity factors measured by batch and elution method as a function of the concentration of ion pairing reagent and methanol can be used to predict the retention in elution method on the basis of capacity factors measured by batch method.

• Korean Journal of Microbiology
• /
• v.33 no.1
• /
• pp.1-6
• /
• 1997

5-Enolpyruvylshikimate 3-phosphate(EPSP) synthase is the sixth enzyme of the shikimate pathway that synthesizes aromatic amino acids. The enzyme is a primary target for the glyphos'lte which is a broad-spectrum and environmetally safe herbicide. As a first step toward development of glyphpsate-resistant EPSP synthase, the EPSP synthase gene(aroA) was amplified by polymerase chain reaction and cluned into pET-25b vector. In this construct. designated pET-aro, the aroA gene is expressed under control of strong T7 promoter. and the EPSP synthase is produced as a fusion protein with pelB leader at N-terminus and HSV-tag and His-tag at C-terminus. When the pET-aro clone was induced to produce the enzyme, it was found that the EPSP synthase was successfully exported to peri plasmic space. The periplasmic transport was greatly dependent on the induction temperatures. Among the induction temperatures examined($25^{\circ}C$, $30^{\circ}C$, $34^{\circ}C$ and $37^{\circ}C$). induction at $34^{\circ}C$ gave rise to maximal periplasmic transport. The recomhinant EPSP synthase could have been purified hy $Ni^{2+}$ -affinity chromatography using the His-tag. and detected hy anti-HSV -tag antibody. The recombinant EPSP synthase also hound to phosphocellulose resin and was eluted hy shikimate 3-phosphate and phosphoenolpyruvate. as expected. The recombinant EPSP synthase purified from phosphocellulose resin showed typical EPSP synthase activity.

• Resources Recycling
• /
• v.31 no.6
• /
• pp.25-35
• /
• 2022

The adsorption/desorption behavior and separation conditions of vanadium and tungsten ions were investigated using a gel-type anion-exchange resin. In the adsorption experiment with the initial acidity of the solution, the adsorption rate of vanadium was remarkably low in strong acids and bases. Additionally, the adsorption rate of tungsten was low in a strong base. An increase in the reaction temperature increased the adsorption reaction rate and maximum adsorption. The effect of tungsten on the maximum adsorption was minimal. The adsorption isotherms of vanadium and tungsten on the ion-exchange resin were suitable for the Langmuir adsorption isotherms of both the ions. For tungsten, the adsorption isotherms of vanadium and tungsten were polyoxometalate. Both ion-exchange resins were simulated using similar quadratic reaction rate models. Vanadium was desorbed in the aqueous solutions of HCl or NaOH, the desorption characteristics of vanadium and tungsten depended on the desorption solution, and tungsten was desorbed in the aqueous solution of NaOH. It was possible to separate the two ions using the desorption process. The desorption reaction reached equilibrium within 30 min, and more than 90% recovery was possible.

• International Journal of Industrial Entomology and Biomaterials
• /
• v.17 no.1
• /
• pp.137-141
• /
• 2008

We previously isolated and cloned a cDNA of abaecin from the Bombus ignitus. In an effort to produce a large amount of soluble abaecin at low cost, we successfully expressed the peptide in Escherichia coli that are highly sensitive to its mature form. For this, we fused the peptide encoding 39 amino acids of mature B. ignitus abaecin to the thioredoxin gene together with a C-terminal 6xHis tag. An enterokinase cleavage site was introduced between the 6xHis tag and mature abaecin to allow final release of the recombinant peptide. A high yield of 9.6 mg soluble fusion protein from 200 ml of bacterial culture was purified by $Ni^{2+}$-charged His-Bind resin affinity column, and 1.4 mg of pure active recombinant abaecin was readily obtained by enterokinase cleavage, followed by affinity chromatograph. The molecular mass of recombinant abaecin peptide was determined by Tricin-SDS-PAGE analysis. The recombinant abaecin exhibited antibacterial activity against Gram-negative bacteria.

• Biotechnology and Bioprocess Engineering:BBE
• /
• v.9 no.5
• /
• pp.362-368
• /
• 2004

A new one-column chromatography process, analogous to a four-zone simulated moving bed (SMB), was presented. The basic principle of the process was identical to that of a four-zone SMB. The process consisted of one chromatographic column and four tanks, instead of the four columns in the four-zone SMB (1-1-1-1), and has been used for the separation of two amino acids, phenylalanine and tryptophan, using an ion exchange resin. The operating parameters for the one-column process and four-zone SMB were obtained from equilibrium theory. Computer simulations were used to compare the performances of the new one column process to that of the general four-zone SMB, using Aspen $Chromatography^{TM}$ v 11.1. The differences between the one-column and SMB processes in terms of the purities and yields of phenylalanine and tryptophan were less than 4 and about $6\%$, respectively. The lower purities of the one-column process were due to the loss of the developed concentration profiles in the column when the liquid was stored in tanks. The one-column process gave great flexibility, and would be useful for reconstructing an existing conventional chromatography process to one of a SMB.

• Biotechnology and Bioprocess Engineering:BBE
• /
• v.10 no.1
• /
• pp.85-90
• /
• 2005

A downstream process was developed for the production of yeast extract from brewer's yeast cells. Various downstream processing conditions including clarification, debittering, and the Maillard reaction were considered in the development of the process. This simple and economic clarification process used flocculating agents, specifically calcium chloride ($1\%$). After the clarification step, a Maillard reaction is initiated as a flavor-enhancing step. By investigating the effects of several operation parameters, including the type of sugar added, sugar dosage, glycine addition, and temperature, on the degree of browning (DB), giucose addition and reaction temperature were found to have significant effects on DB. A synthetic adsorption resin (HP20) was used for the debittering process, which induced a compositional change of the hydrophobic amino acids in the yeast hydrolysate, thereby reducing the bitter taste. The overall dry matter yield and protein yield for the entire process, including the downstream process proposed for the production of brewer's yeast extract were 50 and $50\%$, respectively.

• BMB Reports
• /
• v.37 no.5
• /
• pp.597-602
• /
• 2004

N-terminal His-tagged recombinant $\beta$-1,4-galactosyltransferase from Neisseria meningitidis was expressed and purified to homogeneity by column chromatography using Ni-NTA resin. Mutations were introduced to investigate the roles of, Ser68, His69, Glu88, Asp90, and Tyr156, which are components of a highly conserved region in recombinant $\beta$-1,4 galactosyltransferase. Also, the functions of three other cysteine residues, Cys65, Cys139, and Cys205, were investigated using site-directed mutagenesis to determine the location of the disulfide bond and the role of the sulfhydryl groups. Purified mutant galactosyltransferases, His69Phe, Glu88Gln and Asp90Asn completely shut down wild-type galactosyltransferase activity (1-3%). Also, Ser68Ala showed much lower activity than wild-type galactosyltransferase (19%). However, only the substitution of Tyr156Phe resulted in a slight reduction in galactosyltransferase activity (90%). The enzyme was found to remain active when the cysteine residues at positions 139 and 205 were replaced separately with serine. However, enzyme reactivity was found to be markedly reduced when Cys65 was replaced with serine (27%). These results indicate that conserved amino acids such as Cys65, Ser68, His69, Glu88, and Asp90 may be involved in the binding of substrates or in the catalysis of the galactosyltransferase reaction.

• Journal of the Korean Society of Food Science and Nutrition
• /
• v.29 no.4
• /
• pp.555-560
• /
• 2000

Anthocyanin pigments of purple-fleshed sweet potato (Ipomoea batatas) were extracted with methanol containing 1% HCL and purified with Amberlite IRC-50 cation exchange resin column chromatography. ndividual pigments were isolated by paper chromatography. Among the four bands obtained by paper chromatography, three major bands were identified to be pure pigments by HPLC system. Two pigments were identified through the analysis of acyl moiety, sugar moiety, alkaline degradation products of aglycone, Rf value of paper chromatogram and retention time of HPLC. The anthocyanin pigments of purple-fleshed weet potato seemed to be composed of peonidin-3-diglucoside-5-glucoside acylated with caffeic or ferulic acids.

• Proceedings of the Korean Vacuum Society Conference
• /
• 2013.02a
• /
• pp.110-111
• /
• 2013

FcBP consisting of 13 amino acids specifically binds to Immunoglobulin G Fc domain. Initially, we utilized this peptide for preparation of antibody chip as a PEG composite for enhanced solubility. After then, the peptide conjugate was immobilized on agarose resin, resulting in highly efficient affinity column for antibody purification. The efficiency was comparable to commercial Protein A column. Recently, this peptide was conjugated with cell penetrating peptide (CPP) on a backbone of GFP, affording antibody transducer, which carries antibody into live cells by simple mixing of antibody and the transducer in cell culture media. Antibody transduction into cells was monitored by live cell imaging. More recently, the FcBP was fused to ferritin cage, which consists of 24 ferritin protein molecules. The FcBP-ferritin cage showed greatly increased binding affinity to human IgG. Its binding was analyzed by QCM and SPR analysis. Finally, it was selectively delivered by Herceptin to SKBR3, a breast cancer cell, over MCF10A, non-tumorigenic cells (Fig. 1). Fig. 1. Fluorescent microscopic images of SKBR3 breast cancer cells (A~C) and MCF10A breast cells (D~F) treated with Cy3-trastuzumab/fFcBP-Pf_Fn complexes. Trastuzumab and FcBP-Pf_Fn, which were labeled with Cy3 (Cy3-trastuzumab) and fluorescein (fFcBP-Pf_Fn), respectively, selectively targeted SKBR3 over MCF10A.