• Fisheries and Aquatic Sciences
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• 제26권3호
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• pp.204-215
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• 2023

The thermolabile haemolysin (tlh) of Vibrio parahaemolyticus (Vptlh) from V. parahaemolyticus is a multiple-function enzyme, initially describes as a haemolytic factor activated by lecithin and phospholipase A2 enzymatic activity (Shinoda, 1991; Vazquez-Morado, 2021; Yanagase et al., 1970). Until now, the tlh structure has hypothesized including N-terminal and C-terminal domain, but what domain of the Vptlh structure does the haemolytic activity has not been refined yet. In this study, a 450-bp VpTLH nucleotide sequence of the entire Vptlh gene encoded the C-terminal domain cloned firstly to examine its responsibility in the activity of the Vptlh. The C-terminal domain fused with a 6-His-tag named the His-tag-VpC-terminal domain was expressed successfully in soluble form in the BL21 (DE3) PlysS cell. Remarkably, both expression and purification results confirmed a high agreement in the molecular weight of the His-tag-VpC-terminal domain was 47 kDa. This work showed the His-tag-VpC-terminal domain lysed the erythrocyte membranes in the blood agar and the phosphate buffered saline (0.9%) media without adding the lecithin substrate of the phospholipase enzyme. Haemolysis occurred at all tested diluted concentrations of His-tag-VpC-terminal domain (p < 0.05), providing evidence for the independent haemolytic activity of the His-tag-VpC-terminal domain. The content of 100 ㎍ of the His-tag-VpC-terminal domain brought the highest haemolytic activity of 80% compared to that in the three remaining contents. Significantly, the His-tag-VpC-terminal domain demonstrated not to involve the phospholipase activity in Luria-Bertani agar supplemented with 1% (vol/vol) egg yolk emulsion. All results proved the vital responsibility of the His-tag-VpC-terminal domain in causing the haemolytic activity without the required activation by the phospholipase enzyme. Raw extracts of Phellinus igniarus and Phellinus pipi at 10^-1 mg/mL inhibited the haemolytic activity of the His-tag-VpC-terminal domain from 67.7% to 87.42%, respectively. Hence applying the His-tag-VpC-terminal domain as a simple biological material to evaluate quickly potential derivatives against the Vptlh in vivo conditions will accessible and more advantageous than using the whole of the Vptlh.

• 생명과학회지
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• 제17권9호통권89호
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• pp.1303-1307
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• 2007

A tumor suppressor, merlin is a member of ERM family proteins. It consists of N-terminal FERM domain, ${\alpha}-helical$ region, and C-terminal domain. Alternative splicing of merlin's mRNA generates two isotypes of merlin. Isotype I, which has exon17 at the C-terminus instead of exon16 in isotype II, is known to have tumor suppressor activity. Like other ERM proteins, the C-terminal domain of merlin isotype I interacts to its FERM domain. That of isotype II, however, was reported not to bind FERM domain despite the large common part of C-terminal domain, which possibly binds FERM domain. Here, we show the binding of FERM domain to both C-terminal domains of merlin's two isotypes by isothermal titration calorimetry. These results support that merlin isotype II also can form a closed conformation or a multimer by intramolecular or intermolecular interactions using their FERM domain and C-terminal domain.

• BMB Reports
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• 제33권6호
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• pp.519-524
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• 2000

NF1 proteins are a family of DNA binding proteins which consist of two separate domains, N-terminal DNA binding domain and C-terminal transcription activation domain. The N-terminal 220 amino acids are highly conserved and are also known to mediate dimerization of NF1 proteins. The C-terminal regions of different type of NF1 proteins are heterogeneous and responsible for transcriptional activation. In this study, we tested the interaction between different domains of rat NF1-A protein by yeast two hybrid analysis and observed the interaction between C-terminal regions of NF1-A which do not contain the N-terminal dimerization domain. Our results showed that the C-terminal region of rat NF1-A between residues 231 and 509 strongly interacted not only with itself, but also with human NF1/CTF1 which is a different type of NF1. When the C-terminal region was divided into two fragments, one from residue 231 to 447 and the other from 448 to 509, the two fragments were able to interact with the C-terminal region of NF1-A significantly. This indicates that both fragments contain independent interaction domains. Analysis of the interactions with alanine substituted fragments showed that substitutions of the heptasequence, SPTSPTY of NF1-A, affected interaction between NF1 proteins. Our results strongly suggest that C-terminal regions may also be important for the formation of homo- and heterodimers in addition to the N-terminal dimerization domain. Also, the heptasequence motif may play some roles in dimer formation.

• 대한의생명과학회지
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• 제9권2호
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• pp.105-110
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• 2003

Ankyrins are a ubiquitously expressed family of intracellular adaptor proteins involved in targeting diverse proteins to specialized membrane domains in both the plasma membrane and the endoplasmic reticulum. Canonical ankyrins are 190-220 kDa proteins expressed in most tissues and cell types and comprise a membrane-binding domain (MBD) of 24 ANK repeats, a spectrin-binding domain, a death domain and a C-terminal domain. Rescue studies with ankyrin-B/G chimeras have identified the C-terminal domain of ankyrin-B as the defining domain in specifying ankyrin-B activity, but the function of C-terminal domain of ankyrin-B is, however, not known. We report here that the C-terminal domain of ankyrin-B is capable of interacting with the C-terminal Region of Hsp40. The Hsps are induced not only by heat shock but also by various other environmental stresses. Hsps are also expressed constitutively at normal growth temperatures and have basic and indispensable functions in the life cycle of proteins as molecular chaperones, as well as playing a role in protecting cells from the deleterious stresses. The binding sites required in the interaction between C-terminal domain of ankyrin-B and C-terminal region of Hsp40 were characterized using the yeast two-hybrid system and GST-pull down assay. The interaction between ankyrin-B and Hsp40 represents the first direct evidence of ankyrin's role as chaperones.

• BMB Reports
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• 제52권5호
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• pp.330-335
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• 2019

Hepatitis B virus (HBV) encoding the HBV x protein (HBx) is a known causative agent of hepatocellular carcinoma (HCC). Its pathogenic activities in HCC include interference with several signaling pathways associated with cell proliferation and apoptosis. Mutant C-terminal-truncated HBx isoforms are frequently found in human HCC and have been shown to enhance proliferation and invasiveness leading to HCC malignancy. We investigated the molecular mechanism of the reduced doxorubicin cytotoxicity by C-terminal truncated HBx. Cells transfected with C-terminal truncated HBx exhibited reduced cytotoxicity to doxorubicin compared to those transfected with full-length HBx. The doxorubicin resistance of cells expressing C-terminal truncated HBx correlated with upregulation of the ATP binding cassette subfamily B member 1(ABCB1) transporter, resulting in the enhanced efflux of doxorubicin. Inhibiting the activity of ABCB1 and silencing ABCB1 expression by small interfering ribonucleic acid (siRNA) increased the cytotoxicity of doxorubicin. These results indicate that elevated ABCB1 expression induced by C-terminal truncation of HBx was responsible for doxorubicin resistance in HCC. Hence, co-treatment with an ABCB1 inhibitor and an anticancer agent may be effective for the treatment of patients with liver cancer containing the C-terminal truncated HBx.

• BMB Reports
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• 제56권3호
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• pp.172-177
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• 2023

BEST family is a class of Ca²⁺-activated Cl- channels evolutionary well conserved from bacteria to human. The human BEST paralogs (BEST1-BEST4) share significant amino acid sequence homology in the N-terminal region, which forms the transmembrane helicases and contains the direct calcium-binding site, Ca²⁺-clasp. But the cytosolic C-terminal region is less conserved in the paralogs. Interestingly, this domain-specific sequence conservation is also found in the BEST1 orthologs. However, the functional role of the C-terminal region in the BEST channels is still poorly understood. Thus, we aimed to understand the functional role of the C-terminal region in the human and mouse BEST1 channels by using electrophysiological recordings. We found that the calcium-dependent activation of BEST1 channels can be modulated by the C-terminal region. The C-terminal deletion hBEST1 reduced the Ca²⁺-dependent current activation and the hBEST1-mBEST1 chimera showed a significantly reduced calcium sensitivity to hBEST1 in the HEK293 cells. And the C-terminal domain could regulate cellular expression and plasma membrane targeting of BEST1 channels. Our results can provide a basis for understanding the C-terminal roles in the structure-function of BEST family proteins.

• Bulletin of the Korean Chemical Society
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• 제32권4호
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• pp.1183-1186
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• 2011

The unique Br signature was utilized for C-terminal amino acid sequencing of model peptides. C-terminal carboxyl group was selectively derivatized in peptides, containing side chain carboxyl group, using 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC) and Br was introduced using 4-bromophenylhydrazine hydrochloride (BPH) in a one pot reaction. Matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) tandem mass spectra were obtained carrying the Br signature in the y-series ions. The Br signature facilitated C-terminal sequencing and discrimination of C-terminal carboxyl groups in the free acid and amide forms.

• 한국항해학회지
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• 제20권2호
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• pp.77-84
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• 1996

This is the first report about the development of INMARSAT-C Satellite communication terminal. We analyze the existing Inmarsat-C terminal and examine each rules(IMO rule, domestic rules) about terminal. With that result, we design the basic hardware and software of terminal. This report consists of ; 1) the contents of the overall of operating situation and resources of INMARSAT-C system as like operation of communication system, communication channels and services 2) the contents of the specification of Inmarsat-C terminal hardware and software and the rules of IMO and Type approval 3) the design of basic hardware and reserch of signal modulation/demodulation using Viterbi algorithm 4) the design of software algorithms and functions focused in korean situations.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2001년도 추계학술발표대회
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• pp.727-730
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• 2001

Xanthomonas oryzae #5로부터 클로닝 된 CFTase 의 functional domain의 분석을 위해 CFTase의 recombinant deletion mutant를 구성하고, recombinant protein을 분리, 정제하였다. 분리, 정제한 recombinant protein의 활성을 측정한 결과 C-terminal이 deletion 된 mutant는 cyclization 반응이 소실 되었다. 이와 같은 결과로부터 CFTase의 C-terminal 은 cyclization 반응의 중요한 functional domain 이다.

• Journal of Microbiology and Biotechnology
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• 제20권8호
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• pp.1192-1195
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• 2010

To investigate the influence of the N- or C-terminal regions of pleurocidin (Ple) peptide on antifungal activity, four analogs partially truncated in the N- or C-terminal regions were designed and synthesized. Circular dichroism (CD) spectroscopy demonstrated that all the analogs maintained an alpha-helical structure. The antifungal susceptibility testing also showed that the analogs exhibited antifungal activities against human fungal pathogens, without hemolytic effects against human erythrocytes. The result further indicated that the analogs had discrepant antifungal activities [Ple>Ple (1-22)>Ple (4-25)>Ple (1- 19)>Ple (7-25)] and that N-terminal deletion affected the activities much more than C-terminal deletion. Hydrophobicity [Ple>Ple (1-22)>Ple (4-25)>Ple (1-19)> Ple (7-25)] was thought to have been one of the consistent factors that influenced these activity patterns, rather than the other primary factors like the helicity [Ple>Ple (4-25) >Ple (1-22)>Ple (1-19)>Ple (7-25)] or the net charge [Ple=Ple (4-25)=Ple (7-25)>Ple (1-22)=Ple (1-19)] of the peptides. In conclusion, the hydrophobic amino acids in the N-terminal region of Ple is more crucial for antifungal activity than those in the C-terminal region.