title: Detection of type I collagen fibrils formation and dissociation by a fluorescence method based on thioflavin T.
abstract: In this study, fibrillogenesis and thermal dissociation of pepsin-soluble collagen (PSC), extracted from snakehead (Channa argus) skin, were monitored by fluorescence method based on thioflavin T (Th-T), where the accuracy and sensitivity were evaluated and compared with those of turbidity assay. The fluorescence method revealed the fibrillogenesis dynamics of collagen with better sensitivity, especially at nucleation and plateau stages. The melting temperature (Tm) of PSC was estimated to be 47°C by circular dichroism spectroscopy; below this temperature, the triple-helical structure should be intact. After that, the dynamic process of collagen dissociation was explored by the fluorescence method, and verified by morphological analysis of the fibrils and the proportion of retained fibrils. The thermal dissociation critical temperature (TDCT) of PSC fibrils was confirmed to be 39°C. The fluorescence intensity of fibril-incorporated Th-T gradually decreases in the dissociation process, and the decrease rate can be accelerated by increasing temperature. Finally, the thermal stability of triple-helical structures of free-, assembled- and dissociated-PSC was compared. Thus, we demonstrated the formation and thermal dissociation of collagen fibrils in vitro by a fluorescence method based on Th-T. This approach may advance the understanding of fibril formation and inverse dissociation of fish-sourced collagen in vitro.
authors: School of Food Science and Engineering, Wuhan Polytechnic University, Wuhan, Hubei, China.
authors: Meilian
authors: M
authors: Zou
authors: School of Food Science and Engineering, Wuhan Polytechnic University, Wuhan, Hubei, China.
authors: Huan
authors: H
authors: Yang
authors: School of Chemical and Environmental Engineering, Wuhan Polytechnic University, Wuhan, Hubei, China. Electronic address: wanghaibo@whpu.edu.cn.
authors: Haibo
authors: H
authors: Wang
authors: Tongji medical college, Huzhong University of science and technology, Wuhan, Hubei, China.
authors: Haiyin
authors: H
authors: Wang
authors: School of Chemical and Environmental Engineering, Wuhan Polytechnic University, Wuhan, Hubei, China. Electronic address: zjtwhpu@163.com.
authors: Juntao
authors: J
authors: Zhang
authors: School of Chemical and Environmental Engineering, Wuhan Polytechnic University, Wuhan, Hubei, China.
authors: Benmei
authors: B
authors: Wei
authors: School of Food Science and Engineering, Wuhan Polytechnic University, Wuhan, Hubei, China.
authors: Hanjun
authors: H
authors: Zhang
authors: School of Chemical and Environmental Engineering, Wuhan Polytechnic University, Wuhan, Hubei, China.
authors: Dong
authors: D
authors: Xie
mesh_terms: D000818:Animals; D052160:Benzothiazoles; D024042:Collagen Type I / Q000737:chemistry* / Q000302:isolation & purification; D005109:Extracellular Matrix / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000737:chemistry; D007700:Kinetics; D010434:Pepsin A / Q000737:chemistry; D010473:Perciformes; D044367:Phase Transition; D055550:Protein Stability; D017433:Protein Structure, Secondary; D012867:Skin / Q000737:chemistry*; D012995:Solubility; D012996:Solutions; D013050:Spectrometry, Fluorescence; D013696:Temperature; D013844:Thiazoles / Q000737:chemistry
chemical_list: D052160:Benzothiazoles; D024042:Collagen Type I; D005456:Fluorescent Dyes; D012996:Solutions; D013844:Thiazoles; C009462:thioflavin T; D010434:Pepsin A
keywords: Collagen; Fibrillogenesis; Thioflavin T