• Fisheries and Aquatic Sciences
• /
• 제19권2호
• /
• pp.11.1-11.7
• /
• 2016

In this study, purified peptides from shrimp waste hydrolysates (SWHs) were examined for their inhibitory effects against ${\beta}$-secretase. During consecutive purification using a Sephadex G-25 column chromatography and high performance liquid chromatography on a C18 column, a potent ${\beta}$-secretase inhibitory peptide Asp-Val-Leu-Phe-His (629 Da) was isolated and identified from SWH24 by Q-TOF MS/MS and the $IC_{50}$ value was determined to be $92.70{\mu}M$. The ${\beta}$-secretase inhibition patterns of the purified peptides were found to be competitive. Among synthesized ${\beta}$-secretase inhibitory peptides, Leu-Phe-His had higher ${\beta}$-secretase inhibitory activity than the others. The result of this study suggests that the ${\beta}$-secretase inhibitory peptide derived from SWH24 could be potential candidates to develop nutraceuticals and pharmaceuticals.

• IMMUNE NETWORK
• /
• 제3권2호
• /
• pp.126-135
• /
• 2003

Background: One of the important factors in the prognosis of chronic hepatitis B patient is the degree of replication of hepatitis B virus (HBV). It has been known that HBV DNA polymerase plays the essential role in the replication of HBV. HBV DNA polymerase is composed of four domains, TP (Terminal protein), spacer, RT (Reverse transcriptase) and RNaseH. Among these domains, tyrosine, the $65^{th}$ residue of TP is an important residue in protein-priming reaction that initiates reverse transcription. If monoclonal antibody that recognizes around tyrosine residue were selected, it could be applied to further study of HBV replication. Methods: To produce TP-specific scFv (single-chain Fv) by phage display, mice were immunized using synthetic TP-peptide contains $57{\sim}80^{th}$ amino acid residues of TP domain. After isolation of mRNA of heavy-variable region ($V_H$) and light-chain variable region ($V_L$) from the spleen of the immunized mouse, DNA of $V_H$ and $V_L$ were obtained by RT-PCR and joined by a DNA linker encoding peptide (Gly4Ser)3 as a scFv DNA fragments. ScFv DNA fragments were cloned into a phagemid vector. scFv was expressed in E.coli TG1 as a fusion protein with E tag and phage gIII. To select the scFv that has specific affinity to TP-peptide from the phage-antibody library, we used two cycles of panning and colony lift assay. Results: The TP-peptide-specific scFv was isolated by selection process using TP-peptide as an antigen. Selected scFv had 30 kDa of protein size and its nucleotide sequences were analyzed. Indirect- and competitive-ELISA revealed that the selected scFv specifically recognized both TP-peptide and the HBV DNA polymerase. Conclusion: The scFv that recognizes the TP domain of the HBV DNA polymerase was isolated by phage display.

• 한국환경과학회지
• /
• 제22권12호
• /
• pp.1625-1631
• /
• 2013

In manufacturing of flatfish skin collagen peptide (FSCP) and flatfish protein hydrolysate (FPH) by reuse of dead flatfish from fish farm in Jeju island, the industrial process was optimized with the laboratory scale research and the on-field process. Segmented unit processes from raw material incoming to shipment were established to produce commercial product of FSCP and FPH. Total plate counts of FSCP were twenty five times of FPH, but food poisoning bacteria were not detected in two samples. FSCP and FPH were safe from heavy metal such as Pb(II), Cd(II) and Hg(II). The residual contents of antibiotics and disinfection matter in FSCP and FPH were not detected. The optimized process for mass production made the one-third of the running time and two times of the yield. From economic analysis, the production cost was estimated to 22,000 and 12,000 won/kg for FSCP and FPH, respectively. Therefore the product from the reuse of dead flatfish was expected to have a considerable competitive price and high added-value functional food material compared with other commercially available fish products.

• Journal of Microbiology and Biotechnology
• /
• 제6권1호
• /
• pp.7-11
• /
• 1996

MR-387Al (ARPA-Val-Pro) and A2 (AHPA-Val-Hyp) were prepared as aminopeptidase N inhibitors through the synthesis of peptide MR-387A and B analogues which contained 3-amino-2-hydroxy-4-phenyl butanoic acid (ARPA) as a zinc-chelating moiety. They are competitive inhibitors of aminopeptidase N with inhibition constants(Ki) of 4.1 $\times 10^{-7}\;and 1.1 \times 10^{-6}$ M, respectively. MR-387Al also strongly inhibited aminopeptidase B of human myelogenous leukemia K-562 cell with $IC_50$ of 0.35 $\mu$ M. Inhibitions of aminopeptidase N activity by ARPA-bearing inhibitors of various peptide chain lengths also have been studied. $IC_ 50$ values of AHPA-Val (bestatin), ARPA-Val-Pro (MR-387Al) and ARPA-Val-Pro-Leu (MR-387C) compared against porcine kidney aminopeptidase N were 20.1, 0.60 and 0.08 $\mu$ M, respectively. These results support that a multiple interaction between the $S_1\to S'_3$ sites of aminopeptidase N and the $P_1\to P'_3$ of the inhibitor plays a crucial role in stabilizing strongly the enzyme-inhibitor complex.

• 한국식품영양과학회지
• /
• 제29권4호
• /
• pp.719-725
• /
• 2000

본 연구는 70여종의 국내산 해조류중 가장 높은 ACE 저해 활성을 보였던 김(Porphyra yezoensis, 서천)의 산가수분해물로부터 ACE 활성 저해 펩타이 드를 분리하여 그 특성을 조사하였다. ACE 저해물질의 분획은 균일하게 파쇄한 김을 2.5 N HCl로 산 가수분해한 후 중화하여 한외여과로 분자크기 3 kDa 이하의 물질로 분리하였다. 분자크기 3 kDa이하의 물질에 대하여 column chromatography(Amberlite XAD 8, DEAE-Toyopearl, Sephadex LH-20)와 reverse phase HPLC(C18)를 순차적으로 수행하여 ACE 저해제인 PY3--II-b-h5물질을 분리하였다. PY30-II-b-h5는 분자크기는 약 580 dalton으로 glycine(24.5%), arginine(56.8%), proline(18.8%)의 아미노산 조성을 갖는 저분자 펩타이드였으며, ACE의 저해양상은 경쟁적 저해작용을 하였고, IC50 값은 10.6$\mu\textrm{g}$/mL 이었다.

• 방사선산업학회지
• /
• 제7권2_3호
• /
• pp.191-200
• /
• 2013

In this study, a novel bombesin (BBN) analogues, DOTA-Ala($SO_3H$)-4 aminobenzoyl-Gln-Trp-Ala-Val-Gly-His-Leu-Met-$NH_2$ (DOTA-sBBN) and DOTA-Lys(glucose)-4 aminobenzoyl-Gln-Trp-Ala-Val-Gly-His-Leu-Met-$NH_2$ (DOTA-gluBBN), were synthesized and radiolabeled, and their binding affinities were evaluated. Peptides were prepared by a solid phase synthesis method and their purities were over 98%. DOTA is the chelating agent for $^{177}Lu$-labeling, and the DOTA-conjugated peptides were radiolabeled with $^{177}Lu$ by a high radiolabeling yield (>98%). The Log P values of DOTA-sBBN and DOTA-gluBBN were -2.20 and -2.79, respectively. 50.41% of $^{177}Lu$-DOTA-sBBN and 72.97% of $^{177}Lu$-DOTA-gluBBN were left undegraded by the serum incubation at $37^{\circ}C$ for 48 hr. A competitive displacement of $^{125}I-[Tyr^4]$-BBN on the PC-3 human prostate carcinoma cells revealed that 50% inhibitory concentration ($IC_{50}$) were 1.46 nM of DOTA-sBBN and 4.67 nM of DOTA-gluBBN indicating a highly nanomolar binding affinity for GRPR. Therefore, it is concluded that $^{177}Lu$-DOTA-sBBN and $^{177}Lu$-DOTA-gluBBN can be potential candidates as a targeting modality for the Gastrin-releasing peptide receptor (GRPR)-over-expressing tumors, and further studies to evaluate their biological and pharmacological characteristics are needed.

• 약학회지
• /
• 제38권3호
• /
• pp.351-357
• /
• 1994

The levels of maternal immunity enhancing factor(MIEF), which is an immunomodulatory protein identified from bovine colostrum, were determined by indirect competitive enzyme-linked immunosorbent assay(ELISA) for the colostrum and normal milk collected during the first two weeks of lactation. The mean concentration of MIEF in the colostrum of the first day of lactation was $109\;{\mu}g/ml$, and fell from the third day of lactation to $3{\sim}4\;{\mu}g/ml$. The molecular weight of the purified MIEF determined by reducing SDS-PAGE and TSK G2000SW column chromatography was 22,000 and 24,000 daltons, respectively, showing that MIEF is a monomeric peptide in its native form. To examine the capacity of MIEF to induce differentiation of B Lymphocytes, human tonsillar Iymphocytes were cultured in the presence of different concentrations of MIEF, and then antibody secreting cells were enumerated by enzyme-linked immunospot(ELISPOT) assay. When added to cultures of human tonsillar Lymphocytes, MIEF induced differentiation of resting B Iymphocyte to antibody secreting plasma cells as efficiently as LPS.

• 생명과학회지
• /
• 제27권11호
• /
• pp.1324-1330
• /
• 2017

최근 염분의 섭취와 관련하여 건강에 대한 우려가 늘어남에 따라, 염미대체제에 대한 소비자들의 관심이 급증하고 있으며, 세계적으로 경쟁력 있는 천연 저염제품의 개발이 필요한 실정이다. 최근 연구를 통해 식물성 및 동물성 원료 배합을 통해 가수분해물을 조제하여 최적 화합물을 얻는 것에 성공하였다. 본 연구에서는 염미성 펩타이드 분말내 arginine을 함유한 염미성 펩타이드를 규명하고 정량 하고자 하였다. L-arginine 또는 arginine을 포함한 펩타이드를 표준물질로 하여 $^{13}C$-NMR로 분석한 결과 유사한 위치에 구아니딘기 탄소가 시그널(L-arginine: 156.8 ppm, Arg-Ala: 156.4 ppm, Arg-Ser: 156.4 ppm)이 나타남을 확인하였고, 이를 통해 간편하고 신속하게 arginine 함유 펩타이드 정량분석이 가능할 것으로 기대된다.

• 한국응용약물학회:학술대회논문집
• /
• 한국응용약물학회 1994년도 춘계학술대회 and 제3회 신약개발 연구발표회
• /
• pp.255-255
• /
• 1994

MIEF는 소의 초유중에 함유되어 있는 분자량 22,000 dalton의 peptide로서 말초혈액 림파구의 배양액에 첨가할 경우 자연살해세포를 활성화 시켜 암세포에 대한 자연살해능을 증가시키는 것으로 이미 보고된 바 있다. 본 연구에서는 수유기간별 MIEF의 함량 및 MIEF가 B세포의 증식 및 분화에 미치는 영향을 조사하였다. 수유기간별 MlEF의 함량을 competitive ELISA inhibition assay를 이용하여 조사한 결과 MIEF의 함량은 수유 첫째날의 초유에 109$\mu\textrm{g}$/ml로 가장 높았으며, 그 이후로 급격히 감소하여 3일째 이후에서는 3-4$\mu\textrm{g}$/ml이하로 존재하였다. MIEF가 B세포의 분화에 미치는 영향은 사람의 편도선에서 분리한 림파구의 배양액에 MIEF를 가하고 5-9일간 배양한후 항체를 생산하는 세포의 수를 ELISPOT assay로 측정하여 조사하였다. MIEF는 10-30$\mu\textrm{g}$/ml의 농도에서 B세포의 분화를 촉진시켰으며, 그 정도는 양성대조군으로 사용한 LPS와 유사하여 MIEF가 B세포의 분화를 유도하는 작용이 아주 강력함을 알수 있었다.

• Environmental Analysis Health and Toxicology
• /
• 제12권3_4호
• /
• pp.15-22
• /
• 1997

This experiment was performed to examine the immuno-reactive characteristics of fish metal-binding protein, metallothionein (MT), and gain the practical understandings for the proposed use of fish MT as a biomarker. Liver MT induced by Cd in the silver carp was seperated and purified by gel filtration chromatography and ion exchange chromatography. The immuno-reactivity of fish MT was examined with 3 rabbit antisera. Fish MT showed little reactivity with rabbit anti-rat MT antiserum and a weak reactivity with anti-MT peptide antiserum while showed a strong reactivity with rabbit anti-fish MT antiserum. The time-course change of liver MT in the silver carp, after waterborne exposure to 1 ppm of Cd, was checked by Cd-hem method and established competitive ELISA. In both cases, the induction of liver MT showed a good increasing relationship with the exposure days. The results indicate that the fish MT can be developed as a useful biomonitoring means in the toxicological study and for the evaluation of water pollution.