Putative sequences on Ro60 three-dimensional structure accessible for 4-hydroxy-2-nonenal (HNE) modification compared to in vitro HNE modification of Ro60 sequences

It is difficult to a produce highly functional bioartificial liver (BAL) using only hepatocytes, because it is believed that liver-specific three-dimensional structure is necessary to maintain high function for BAL. But it is difficult to construct a culture system with liver-specific three-dimensional structure in vitro. To realize a highly functional culture system with liver-specific three-dimensional structure, we developed a culture system using liver slices that keep liver-specific architecture, such as liver lobule and hepatic microvascular system. Liver slices were embedded in agarose gel to maintain them under a moist and three-dimensional environment. We examined the viability and function of liver slices by using various shapes of agarose gel. Liver slices were cultured 1) under stationary condition (control), 2) directly embedded in gel, and 3) embedded in cylindrical gel for good drainage of medium and ventilation of air. The viability and function of the incubated liver slices were evaluated by LDH leakage, histomorphology, and immunohistochemistry. At 10 days, the morphological condition and function of liver slices embedded in cylindrical gel were maintained better than liver slices directly embedded in gel or in the stationary condition. We suggest that high functionality and morphological condition of liver slices could be maintained by embedding in cylindrical gel. In the future, it is possible that this method could be used to develop a highly functional bioartificial liver.

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The three-dimensional structure of in vitro reconstituted Xenopus laevis chromosomes by EM tomography

Chromosoma ◽

10.1007/s00412-007-0101-0 ◽

2007 ◽

Vol 116 (4) ◽

pp. 349-372 ◽

Cited By ~ 32

Author(s):

Peter König ◽

Michael B. Braunfeld ◽

John W. Sedat ◽

David A. Agard

Keyword(s):

Xenopus Laevis ◽

Three Dimensional ◽

Dimensional Structure ◽

Three Dimensional Structure ◽

Em Tomography

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NMR three-dimensional structure of the cationic peptide Stigmurin from Tityus stigmurus scorpion venom: In vitro antioxidant and in vivo antibacterial and healing activity

Peptides ◽

10.1016/j.peptides.2020.170478 ◽

2021 ◽

Vol 137 ◽

pp. 170478

Author(s):

Alessandra Daniele-Silva ◽

Suedson de Carvalho Silva Rodrigues ◽

Elizabeth Cristina Gomes dos Santos ◽

Moacir Fernandes de Queiroz Neto ◽

Hugo Alexandre de Oliveira Rocha ◽

...

Keyword(s):

Three Dimensional ◽

Scorpion Venom ◽

Dimensional Structure ◽

Cationic Peptide ◽

Three Dimensional Structure

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Frequent SLC12A3 mutations in Chinese Gitelman syndrome patients: structure and function disorder

Endocrine Connections ◽

10.1530/ec-21-0262 ◽

2021 ◽

Author(s):

Lanping Jiang ◽

Xiaoyan Peng ◽

Bingbin Zhao ◽

Lei Zhang ◽

Lubin Xu ◽

...

Keyword(s):

Three Dimensional ◽

Gene Mutations ◽

Dimensional Structure ◽

Novel Mutations ◽

Three Dimensional Structure ◽

Slc12a3 Gene ◽

Uptake Experiment ◽

Frequent Mutations ◽

22Na Uptake

Purposes: This study was conducted to identify the frequent mutations from reported Chinese Gitelman syndrome (GS) patients, to predict three-dimensional structure change of human Na-Cl co-transporter (hNCC), and to test the activity of these mutations and some novel mutations in vitro and in vivo. Methods: SLC12A3 gene mutations in Chinese GS patients previously reported in the PubMed, CNKI and Wanfang database were summarized. Predicted configurations of wild type (WT) and mutant proteins were achieved using the I-TASSER workplace. Six missense mutations (T60M, L215F, D486N, N534K, Q617R and R928C) were generated by site-directed mutagenesis. 22Na+ uptake experiment was carried out in the Xenopus laevis oocyte expression system. 35 GS patients and 20 healthy volunteers underwent the thiazide test. Results: T60M, T163M，D486N, R913Q, R928C and R959 frameshift were frequent SLC12A3 gene mutations (mutated frequency >3%) in 310 Chinese GS families. The protein’s three-dimensional structure was predicted to be altered in all mutations. Compared with WT hNCC, the thiazide-sensitive 22Na+ uptake was significantly diminished for all 6 mutations: T60M 22±9.2%, R928C 29±12%, L215F 38±14%, N534K 41±15.5%, Q617R 63±22.1% and D486N 77±20.4%. In thiazide test, the net increase in chloride fractional excretion in 20 healthy controls was significantly higher than GS patients with or without T60M or D486N mutations. Conclusions: Frequent mutations (T60M, D486N, R928C) and novel mutations (L215F, N534K and Q617R) lead to protein structure alternation and protein dysfunction verified by 22Na+ uptake experiment in vitro and thiazide test on patients.

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In Vivo and In Vitro Studies of Bacillus subtilis Ferrochelatase Mutants Suggest Substrate Channeling in the Heme Biosynthesis Pathway

Journal of Bacteriology ◽

10.1128/jb.184.14.4018-4024.2002 ◽

2002 ◽

Vol 184 (14) ◽

pp. 4018-4024 ◽

Cited By ~ 22

Author(s):

Ulf Olsson ◽

Annika Billberg ◽

Sara Sjövall ◽

Salam Al-Karadaghi ◽

Mats Hansson

Keyword(s):

Bacillus Subtilis ◽

Amino Acid ◽

Three Dimensional ◽

Site Directed Mutagenesis ◽

Dimensional Structure ◽

Amino Acid Residues ◽

Three Dimensional Structure ◽

Activity Measurements

ABSTRACT Ferrochelatase (EC 4.99.1.1) catalyzes the last reaction in the heme biosynthetic pathway. The enzyme was studied in the bacterium Bacillus subtilis, for which the ferrochelatase three-dimensional structure is known. Two conserved amino acid residues, S54 and Q63, were changed to alanine by site-directed mutagenesis in order to detect any function they might have. The effects of these changes were studied in vivo and in vitro. S54 and Q63 are both located at helix α3. The functional group of S54 points out from the enzyme, while Q63 is located in the interior of the structure. None of these residues interact with any other amino acid residues in the ferrochelatase and their function is not understood from the three-dimensional structure. The exchange S54A, but not Q63A, reduced the growth rate of B. subtilis and resulted in the accumulation of coproporphyrin III in the growth medium. This was in contrast to the in vitro activity measurements with the purified enzymes. The ferrochelatase with the exchange S54A was as active as wild-type ferrochelatase, whereas the exchange Q63A caused a 16-fold reduction in V max. The function of Q63 remains unclear, but it is suggested that S54 is involved in substrate reception or delivery of the enzymatic product.

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Three-Dimensional Structure of Aleutian Mink Disease Parvovirus: Implications for Disease Pathogenicity

Journal of Virology ◽

10.1128/jvi.73.8.6882-6891.1999 ◽

1999 ◽

Vol 73 (8) ◽

pp. 6882-6891 ◽

Cited By ~ 55

Author(s):

Robert McKenna ◽

Norman H. Olson ◽

Paul R. Chipman ◽

Timothy S. Baker ◽

Tim F. Booth ◽

...

Keyword(s):

Capsid Protein ◽

Parvovirus B19 ◽

Three Dimensional ◽

Dimensional Structure ◽

Minute Virus Of Mice ◽

Image Reconstruction Techniques ◽

Three Dimensional Structure ◽

Feline Panleukopenia Virus

ABSTRACT The three-dimensional structure of expressed VP2 capsids of Aleutian mink disease parvovirus strain G (ADVG-VP2) has been determined to 22 Å resolution by cryo-electron microscopy and image reconstruction techniques. A structure-based sequence alignment of the VP2 capsid protein of canine parvovirus (CPV) provided a means to construct an atomic model of the ADVG-VP2 capsid. The ADVG-VP2 reconstruction reveals a capsid structure with a mean external radius of 128 Å and several surface features similar to those found in human parvovirus B19 (B19), CPV, feline panleukopenia virus (FPV), and minute virus of mice (MVM). Dimple-like depressions occur at the icosahedral twofold axes, canyon-like regions encircle the fivefold axes, and spike-like protrusions decorate the threefold axes. These spikes are not present in B19, and they are more prominent in ADV compared to the other parvoviruses owing to the presence of loop insertions which create mounds near the threefold axes. Cylindrical channels along the fivefold axes of CPV, FPV, and MVM, which are surrounded by five symmetry-related β-ribbons, are closed in ADVG-VP2 and B19. Immunoreactive peptides made from segments of the ADVG-VP2 capsid protein map to residues in the mound structures. In vitro tissue tropism and in vivo pathogenic properties of ADV map to residues at the threefold axes and to the wall of the dimples.

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Generation of Glucose-Responsive Functional Islets with a Three-Dimensional Structure from Mouse Fetal Pancreatic Cells and iPS Cells In Vitro

PLoS ONE ◽

10.1371/journal.pone.0028209 ◽

2011 ◽

Vol 6 (12) ◽

pp. e28209 ◽

Cited By ~ 30

Author(s):

Hiroki Saito ◽

Masaki Takeuchi ◽

Kazuhiro Chida ◽

Atsushi Miyajima

Keyword(s):

Three Dimensional ◽

Ips Cells ◽

Dimensional Structure ◽

Three Dimensional Structure ◽

Pancreatic Cells

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Folding of the td pre-RNA with the help of the RNA chaperone StpA

Biochemical Society Transactions ◽

10.1042/bst0301175 ◽

2002 ◽

Vol 30 (6) ◽

pp. 1175-1180 ◽

Cited By ~ 11

Author(s):

O. Mayer ◽

C. Waldsich ◽

R. Grossberger ◽

R. Schroeder

Keyword(s):

Conformational Changes ◽

Three Dimensional ◽

Group I Intron ◽

Dimensional Structure ◽

Rna Chaperone ◽

Three Dimensional Structure ◽

Group I ◽

The Impact

The td group I intron is inserted in the reading frame of the thymidylate synthase gene, which is mainly devoid of structural elements. In vivo, translation of the pre-mRNA is required for efficient folding of the intron into its splicing-competent structure. The ribosome probably resolves exon-intron interactions that interfere with splicing. Uncoupling splicing from translation, by introducing a non-sense codon into the upstream exon, reduces the splicing efficiency of the mutant pre-mRNA. Alternatively to the ribosome, co-expression of genes that encode proteins with RNA chaperone activity promote folding of the td pre-mRNA in vivo. These proteins also efficiently accelerate folding of the td pre-mRNA in vitro. In order to understand the mechanism of action of RNA chaperones, we probed the impact of the RNA chaperone StpA on the structure of the td intron in vivo and compared it with that of the well characterized Cyt-18 protein, which is a group-I-intron-specific splicing factor. We found that the two proteins have opposite effects on the structure of the td intron. While StpA loosens the three-dimensional structure, Cyt-18 tightens it up. Furthermore, mutations that destabilize the intron structure render the mutants sensitive to StpA, whereas splicing of these mutants is rescued by Cyt-18. Our results provide direct evidence for protein-induced conformational changes within a catalytic RNA in vivo. Whereas StpA resolves tertiary contacts enabling the RNA to refold, Cyt-18 contributes to the stabilization of the native three-dimensional structure.

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