• Animal cells and systems
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• 제1권4호
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• pp.589-594
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• 1997

The neuronal voltage-sensitive calcium channels (VSCCs) are multisubunit complexes consisting of $\alpha_1,\;\alpha_2-\delta$ and $\beta$ subunits. Heterologous expression and biochemical studies have shown that the activity of VSCCs is regulated by their $\beta$ subunits in a $\beta$ subunit isoform-specific manner. To elucidate the $\beta$ subunit identity of the P/Q-type calcium channel encoded by an $\alpha_{1A}$ subunit, which is exclusively expressed in the Purkinje and granule cell of the cerebellum, we have examined the spatial and temporal expression patterns of $\beta$ subunits and compared them with those of $\alpha_{1A}$ subunit in the developing rat cerebellum. Reverse transcriptase- polymerase chain reaction (RT-PCR) and Northern blot analysis have shown that $\beta_4$ subunit mRNA was prominently expressed in the cerebellum and much more abundant than any other distinct $\beta$ subunits. RNase protection assay has further demonstrated that the expression of $\alpha_{1A}$ and $\beta_4$ subunits increased during cerebellar development, while the amount of $\beta_2$ and $\beta_3$ mRNAs did not significantly change. In addition, a $\beta_4$ transcript was present in cultured cerebellar granule cells, but not in astrocyte cells, and the level of $\beta_4$ mRNA expression increased gradually in vitro seen as in vivo. Based on the spatial and temporal expression patterns of $\beta_4$ subunit, we conclude that $\beta_4$ may predominantly associate, but probably not exclusively, with the $\alpha_{1A}$ subunit in rat cerebellar granule cells.

• Journal of Cancer Prevention
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• 제23권4호
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• pp.153-161
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• 2018

Imbalance of protein homeostasis (proteostasis) is known to cause cellular malfunction, cell death, and diseases. Elaborate regulation of protein synthesis and degradation is one of the important processes in maintaining normal cellular functions. Protein degradation pathways in eukaryotes are largely divided into proteasome-mediated degradation and lysosome-mediated degradation. Proteasome is a multisubunit complex that selectively degrades 80% to 90% of cellular proteins. Proteasome-mediated degradation can be divided into 26S proteasome (20S proteasome + 19S regulatory particle) and free 20S proteasome degradation. In 1980, it was discovered that during ubiquitination process, wherein ubiquitin binds to a substrate protein in an ATP-dependent manner, ubiquitin acts as a degrading signal to degrade the substrate protein via proteasome. Conversely, 20S proteasome degrades the substrate protein without using ATP or ubiquitin because it recognizes the oxidized and structurally modified hydrophobic patch of the substrate protein. To date, most studies have focused on protein degradation via 26S proteasome. This review describes the 26S/20S proteasomal pathway of protein degradation and discusses the potential of proteasome as therapeutic targets for cancer treatment as well as against diseases caused by abnormalities in the proteolytic system.

• Journal of Plant Biotechnology
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• 제47권4호
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• pp.283-288
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• 2020

Although the evolution of Arabidopsis thaliana and humans diverged approximately 1.6 billion years ago, recent studies have demonstrated that protein function and cellular processes involved in disease response remain remarkably conserved. Particularly, γ-secretase, a multisubunit protein complex that participates in intramembrane proteolysis (RIP) regulation, is also known to mediate the cleavage of more than 80 substrates including the amyloid precursor protein (APP) and the Notch receptor. Although the genes (PS1/2, APH-1, PEN-2, and NCT) coding for the γ-secretase complex components are present in plant genomes, their function remains largely uncharacterized. Given that the deposition of 42 amino acid long amyloid-β peptides (hAβ₄₂) is thought to be one of the main causes of Alzheimer's disease, we aimed to examine the physiological effects of hAβ₄₂ peptides on plants. Interestingly, we found that Arabidopsis protoplast death increased after 24 h of exposure to 3 or 5 µM hAβ₄₂ peptides. Furthermore, transgenic Arabidopsis plants overexpressing the hAβ₄₂ gene exhibited changes in primary root length and silique phyllotaxy. Taken together, our results demonstrate that hAβ₄₂ peptides, a metazoan protein, significantly affect Arabidopsis protoplast viability and plant morphology.

• 미생물학회지
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• 제46권1호
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• pp.96-103
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• 2010

단백질의 N-글리코실화는 대표적인 번역 후 변형 중의 하나로 진핵생물 뿐 아니라 원핵생물에서도 발견된다. N-글리코실화는 단백질 상의 N-글리코실 서열인 N-x-S/T 위치에 지질과 연결된 올리고당(lipid-linked oligosaccharide, LLO)으로부터 올리고당 전이효소(oligosaccharyltransferase, OTase) 활성에 의해 글리칸(glycan)이 전달되어 당단백질의 합성이 이루어진다. 본 연구에서는 OTase의 세포내 활성을 측정하기 위하여 5/6-carboxyltetramethylrhodamine (TAMRA)이 도입된 형광펩타이드 TAMRA-DA$\underline{N}$Y$\underline{T}$K-$NH_2$를 이용하였다. OTase활성 측정은 단일 서브유닛으로 효소의 활성을 갖는 운동핵 편모충류인 Leishmania major Stt3p와 병원성 미생물인 Campylobacter jejuni PglB를 진핵생물과 원핵생물의 모델 효소로 각각 사용하여 Saccharomyces cerevisiae와 C. jejuni 유래 LLO와 형광 펩타이드를 반응시켜 당-펩타이드를 합성하였다. 합성된 당-펩타이드를 미반응한 형광펩타이드와 분리 및 당-펩타이드의 정량 분석을 위하여 Tricine SDS-PAGE, ConA 렉틴 컬럼 및 fluorospectrophotometer, HPLC를 사용하였으며, 당-펩타이드 분석을 통해 각 방법의 장단점을 비교하였다. 비교 분석 결과 Tricine SDS-PAGE를 이용한 형광 이미지 분석과, 렉틴 컬럼을 통해 분리된 당-펩타이드의 fluorospectrophotometer 정량법에 비해, HPLC를 이용한 방법이 OTase에 의해 생성된 당-펩타이드를 분석하는데 더 정확하고 정량적인 값을 제시하는 것으로 확인되었다.