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Received July 31, 2022
Accepted October 8, 2022
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Novel silicatein-like protein for biosilica production from Amphimedon queenslandica and its use in osteogenic composite fabrication
1Department of Biotechnology and Bioinformatics, Korea University, Sejong 30019, Korea 2Institute of Industrial Technology, Korea University, Sejong 30019, Korea 3, Korea 4Department of Botany and Microbiology, Faculty of Science, Minia University, Minia 61519, Egypt
spack@korea.ac.kr
Korean Journal of Chemical Engineering, February 2023, 40(2), 419-428(10), 10.1007/s11814-022-1314-x
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Abstract
Efforts to find sustainable and eco-friendly ways to conduct chemical reactions have led to the mimicking of nature. In this study, a new silica polymerization protein that can produce silica in an environmentally friendly manner was developed using cathepsin L-like protein (AqCtL) from Amphimedon queenslandica with a 61% sequence identity to that of silicatein-alpha, which is a natural biosilicifying enzyme. To stabilize the protein structure, heterologously expressed AqCtL in Escherichia coli was mutated into AqCtLSN by changing the amino acid residues responsible for protease cleavage. The insoluble form of AqCtLSN was reconstituted into a soluble protein through the refolding process, displaying silica-condensing activity from silicic acid. AqCtLSN self-assembled, aggregated, and attached to a support in the PBS buffer without losing silica deposition activity. These properties were applied to fabricate a silica-hybrid material using a gelatin-tyramine-alginate cross-linked hydrogel as a scaffold. FT-IR analysis revealed that a silica hybrid material was produced owing to the in situ silicification by AqCtLSN immobilized on the hydrogel. The surface of biosilica mediated by AqCtLSN demonstrated an increase in cell proliferation, alkaline phosphatase activity, and calcium mineral precipitation in the osteogenesis of MC3T3 E1 cells compared to those without biosilica. In conclusion, AqCtLSN, recombinantly expressed in E. coli, is a novel biosilica-forming protein that can be used to produce composites for biomedical applications, especially bone regeneration.
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Ki MR, Yeo KB, Pack SP, Bioprocess. Biosyst. Eng., 36, 643 (2012)
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Lipinszki Z, Vernyik V, Farago N, Sari T, Puskas LG, Blattner FR, Posfai G, Gyorfy Z, ACS Synth. Biol., 7, 2656 (2018)
Menard R, Carmona E, Takebe S, Dufour E, Plouffe C, Mason P, Mort JS, J. Biol. Chem., 273, 4478 (1998)
Bendtsen JD, Nielsen H, von Heijne G, Brunak S, J. Mol. Biol., 340, 783 (2004)
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Kozlowska J, Stachowiak N, Sionkowska A, Polymers, 10, 456 (2018)
Jiang BP, Zhang L, Zhu Y, Shen XC, Ji SC, Tan XY, Cheng L, Liang H, J. Mater. Chem. B, 3, 3767 (2015)
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Plowright R, Dinjaski N, Zhou S, Belton DY, Kaplan DL, Perry CC, RSC Adv., 6, 21776 (2016)
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Zhu W, Gao X, Zou X, Müller WEG, Wang S, Wang Y, Liu Y, J. Biomater Tissue Eng., 8, 258 (2018)
Dudik O, Amorim S, Xavier JR, Rapp HT, Silva TH, Pires RA, Reis RL, Front. Mar. Sci., 8, 637810 (2021)
