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• 修士

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• Lowering the culture temperature corrects collagen abnormalities caused by HSP47 gene knockout.

  Kazunori K Fujii, Yuki Taga, Takayuki Sakai, Shinya Ito, Shunji Hattori, Kazuhiro Nagata, Takaki Koide

  Scientific reports   9 ( 1 ) 17433 - 17433  2019年11月  [査読有り]  [国際誌]

  　概要を見る

  Heat shock protein 47 (HSP47) is an endoplasmic reticulum (ER)-resident molecular chaperone that specifically recognizes triple helical portions of procollagens. The chaperone function of HSP47 is indispensable in mammals, and hsp47-null mice show an embryonic lethal phenotype accompanied by severe abnormalities in collagen-based tissue structures. Two leading hypotheses are currently accepted for the molecular function of HSP47 as a procollagen-specific chaperone. One is facilitation of procollagen folding by stabilizing thermally unstable triple helical folding intermediates, and the other is inhibition of procollagen aggregation or lateral association in the ER. The aim of this study was to elucidate the functional essence of this unique chaperone using fibroblasts established from hsp47-/- mouse embryos. When the cells were cultured at 37 °C, various defects in procollagen biosynthesis were observed, such as accumulation in the ER, over-modifications including prolyl hydroxylation, lysyl hydroxylation, and further glycosylation, and unusual secretion of type I collagen homotrimer. All defects were corrected by culturing the cells at a lower temperature of 33 °C. These results indicated that lowering the culture temperature compensated for the loss of HSP47. This study elucidated that HSP47 stabilizes the elongating triple helix of procollagens, which is otherwise unstable at the body temperature of mammals.

  DOI PubMed

• Structural optimization of cyclic peptides that efficiently detect denatured collagen.

  Takita KK, Fujii KK, Ishii K, Koide T

  Organic & biomolecular chemistry   17 ( 31 ) 7380 - 7387  2019年08月  [査読有り]

  DOI PubMed

• Cyclic Peptides for Efficient Detection of Collagen

  Koh K. Takita, Kazunori K. Fujii, Tetsuya Kadonosono, Ryo Masuda, Takaki Koide

  ChemBioChem   19 ( 15 ) 1613 - 1617  2018年08月  [査読有り]

  　概要を見る

  © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim We report here a new class of collagen-binding peptides, cyclic collagen-mimetic peptides (cCMPs), that efficiently hybridize with the triple-helix-forming portions of collagen. cCMPs are composed of two parallel collagen-like (Xaa-Yaa-Gly)n strands with both termini tethered by covalent linkages. Enzyme-linked immunosorbent assays and western blotting analysis showed that cCMPs exhibit more potent affinity toward collagen than reported collagen-binding peptides and can specifically detect different collagen polypeptides in a mixture of proteins. Collagen secreted from cultured cells was detected by confocal microscopy with fluorescein-labeled cCMP. The cCMP is also shown to detect sensitively folding intermediates in the endoplasmic reticulum, something that was difficult to visualize with conventional collagen detectors. Molecular-dynamics simulations suggested that a cCMP forms a more stably hybridized product than its single-chain counterpart; this could explain why cCMP has higher affinity toward denatured collagen. These results indicate the usefulness of cCMPs as tools for detecting denatured collagen.

  DOI PubMed

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• 環境温度が魚類コラーゲンに与える影響

  2020年   小出　隆規

  　概要を見る

  本研究では、環境の温度が変温動物である魚類のコラーゲンの性質にどのような影響を与えるのかを解明することを目指している。コラーゲンは3本のポリペプチド鎖からなる3重らせん構造をもつタンパク質であり、その構造は翻訳後修飾などにより熱的に安定化されている。この安定化の度合いが環境の温度によって変化しているのかを明らかにすべく、魚類細胞を複数の異なる温度で培養し、分泌されたコラーゲンの熱変性温度を測定した。その結果、高温で培養した際に分泌されたコラーゲンでは、3重らせん構造の熱変性温度が上昇していることが明らかになった。