• Title/Summary/Keyword: cryo-electron microscopy

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Single-Molecule Imaging Reveals the Mechanism Underlying Histone Loading of Schizosaccharomyces pombe AAA+ ATPase Abo1

  • Kang, Yujin;Cho, Carol;Lee, Kyung Suk;Song, Ji-Joon;Lee, Ja Yil
    • Molecules and Cells
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    • v.44 no.2
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    • pp.79-87
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    • 2021
  • Chromatin dynamics is essential for maintaining genomic integrity and regulating gene expression. Conserved bromodomain-containing AAA+ ATPases play important roles in nucleosome organization as histone chaperones. Recently, the high-resolution cryo-electron microscopy structures of Schizosaccharomyces pombe Abo1 revealed that it forms a hexameric ring and undergoes a conformational change upon ATP hydrolysis. In addition, single-molecule imaging demonstrated that Abo1 loads H3-H4 histones onto DNA in an ATP hydrolysis-dependent manner. However, the molecular mechanism by which Abo1 loads histones remains unknown. Here, we investigated the details concerning Abo1-mediated histone loading onto DNA and the Abo1-DNA interaction using single-molecule imaging techniques and biochemical assays. We show that Abo1 does not load H2A-H2B histones. Interestingly, Abo1 deposits multiple copies of H3-H4 histones as the DNA length increases and requires at least 80 bp DNA. Unexpectedly, Abo1 weakly binds DNA regardless of ATP, and neither histone nor DNA stimulates the ATP hydrolysis activity of Abo1. Based on our results, we propose an allosteric communication model in which the ATP hydrolysis of Abo1 changes the configuration of histones to facilitate their deposition onto DNA.

An Immune-Electron Microscopic Study for Cluster Designation on the Phagocytic Synovial Cells in the Knee Joint of the Human (인체 무릎관절 윤활포식세포 cluster designation 표지에 관한 면역전자현미경적 연구)

  • Lim, Hyoung-Soo;Cho, Kook-Hyeung;Kim, Yong-Wook;Park, Kyeong-Han;Hwang, Young-Il;Chang, Ka-Young;Hwang, Douk-Ho
    • Applied Microscopy
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    • v.30 no.2
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    • pp.173-183
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    • 2000
  • This study was designed to observe the ultrastructural localization of synoviocytes, which are concerned with the function of phagocytic synovial cells (type A synoviocytes, macrophage-like synoviocytes), in the knee joint of the human for CD14 and CD105 by cryo-immune-electron microscopic technique. The synovium were dissected and fixed for two hours (in 4% paraformaldehyde and 0.1% glutaraldehyde mixture), and were immerged in 2.3 M sucrose and 20% PVP solution. Finally, they were cut with the cryoultramicrotome and labelled with primary antibodies (monoclonal mouse anti-human CD14, monoclonal mouse anti-human CD105 (endoglin) and secondary (donkey anti-mouse IgG) tagged with 6 nm colloidal gold particles. The tissues were observed under transmission electron microscope. This study was resulted as follows. 1. In the synovium of the human knee joint, CD14+ cells were identified. These cells showed phagocytic synovial cell's features. In the phagocytic synoviocyte, the distributions of CD14 were marked in the cytoplasm, around vacuoles, and in cytoplasmic process, but not detected inside of vacuoles. 2. In the synovium of the human knee joint, CD105+ cells were identified. These cells were recognized endothelial cells and phagocytic synovial cells. In the phagocytic synovial cells, the distributions of CD105 (endoglin) were marked in cytoplasic process, around vacuoles, and in cell membrane, but not detected inside of vacuoles. On the basis of above findings, it is obvious that phagocytic synovial cells were marked at CD 14 and CD 105, and might be play the role of activated macrophages or phagocytes in the synovial membrane.

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Phase Behavior Study of Fatty Acid Potassium Cream Soaps (지방산 칼륨 Cream Soaps 의 상거동 연구)

  • Noh, Min Joo;Yeo, Hye Lim;Lee, Ji Hyun;Park, Myeong Sam;Lee, Jun Bae;Yoon, Moung Seok
    • Journal of the Society of Cosmetic Scientists of Korea
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    • v.48 no.1
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    • pp.55-64
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    • 2022
  • The potassium cream soap with fatty acid called cleaning foam has a crystal gel structure, and unlike an emulsion system, it is weak to shear stress and shows characteristics that are easily separated under high temperature storage conditions. The crystal gel structure of cleansing foams is significantly influenced by the nature and proportion of fatty acids, degree of neutralization, and the nature and proportion of polyols. In order to investigate the effect of these parameters on the crystal gel structure, a ternary system consisting of water/KOH/fatty acid was investigated in this study. The investigation of differential scanning calorimeter (DSC) revealed that the eutectic point was found at the ratio of myristic acid (MA) : stearic acid (SA) = 3 : 1 and ternary systems were the most stable at the eutectic point. However, the increase in fatty acid content had little effect on stability. On the basis of viscosity and polarized optical microscopy (POM) measurements, the optimum degree of neutralization was found to be about 75%. The system was stable when the melting point (Tm) of the ternary system was higher than the storage temperature and the crystal phase was transferred to lamellar gel phase, but the increase in fatty acid content had little effect on stability. The addition of polyols to the ternary system played an important role in changing the Tm and causing phase transition. The structure of the cleansing foams were confirmed through cryogenic scanning electron microscope (Cryo-SEM), small and wide angle X-ray scattering (SAXS and WAXS) analysis. Since butylene glycol (BG), propylene glycol (PG), and dipropylene glycol (DPG) lowered the Tm and hindered the lamellar gel formation, they were unsuitable for the formation of stable cleansing foam. In contrast, glycerin, PEG-400, and sorbitol increased the Tm, and facilitated the formation of lamellar gel phase, which led to a stable ternary system. Glycerin was found to be the most optimal agent to prepare a cleansing foam with enhanced stability.

A Study on the Formation of Lamellar Liquid Crystalline Using Skin Mimicking Surfactant (피부모사체 계면활성제를 사용한 라멜라 액정의 생성에 관한 연구)

  • Kim, In-Young;Nam, Eun-Hee;Shin, Moon-Sam
    • Journal of the Korean Applied Science and Technology
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    • v.37 no.3
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    • pp.484-495
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    • 2020
  • This study is a mixed surfactant (MimicLipid-MSM1000) that forms the same structure as that of the stratum corneum, sucrose distearate, polyglyceryl-2 dioleate, fermented squalane, ergosterol, 10-hydroxystearic acid, mixture consisting of was synthesized. When using 2~5 wt% of this mixed surfactant, it was possible to make an artificial skin mimetic that forms a multi-layer lamellar structure of 5~30 layers. An emulsion was prepared using this mixed surfactant, and a multi-layered lamellar phase was formed and analyzed mechanically. The appearance of this surfactant was a light brown hard wax, the hydrophilic lipophilic balance (HLB) was 12.53, the critical parameter value was 0.987, and the acid value was 0.13. Stability according to pH change was also stable in acidic (3.8), neutral (7.2) and alkaline (10.8). The particle size of the liquid crystal was found to be the most stable maltese cross lamellar crystalline droplet at 5~25mm. The size of the emulsified particles according to the change in the speed of the homo agitator is 2500 rpm (17.9mm±2.6mm), 3500rpm (12.5mm±2.1mm), 4500rpm (6.2mm±1.8mm) particles were formed. Liquid crystal forming particles were observed through a polarization microscope, and the formation structure of the liquid crystal was precisely analyzed with a scanning electron microscope (cryo-SEM). As an application field, it is expected that it will be widely applicable to the development of various prescriptions, such as various skin care cosmetics, makeup care cosmetics, and scalp protection cosmetics, by using a skin-mimicking surfactant.