TY - JOUR
T1 - Dual-Functionalizable Streptavidin-SpyCatcher-Fused Protein-Polymer Hydrogels as Scaffolds for Cell Culture
AU - Minamihata, Kosuke
AU - Hamada, Yusei
AU - Kagawa, Genki
AU - Ramadhan, Wahyu
AU - Higuchi, Ayato
AU - Moriyama, Kousuke
AU - Wakabayashi, Rie
AU - Goto, Masahiro
AU - Kamiya, Noriho
N1 - Funding Information:
This work was supported by the Japan Society for the Promotion of Science, JSPS KAKENHI (grant numbers JP20K05234 to K.M. and JP19H00841 to N.K.). Victoria Muir, PhD, from Edanz Group ( https://en-author-services.edanzgroup.com/ac ) edited a draft of this manuscript. We thank Dr. Yunmei Mu and Assoc. Prof. Takeshi Mori in Kyushu University for the support of conducting qPCR analysis.
Publisher Copyright:
©
PY - 2020/11/16
Y1 - 2020/11/16
N2 - Hydrogels possessing the ability to control cell functions have great potential as artificial substrates for cell culture. Herein, we report dual-functionalizable protein-polymer hybrid hydrogels prepared by thiol oxidation catalyzed by horseradish peroxidase and a phenolic molecule. A chimera protein of streptavidin (SA) and the SpyCatcher protein, with a cysteine residue at its N-terminus, (C-SA-SC) was constructed and co-cross-linked with thiol-functionalized four-arm polyethylene glycol (PEG-SH) to obtain hydrogels possessing two orthogonal conjugation moieties. Hydrogel formation using C-SA-SC conjugated with biotinylated or SpyTagged functional molecules (premodification strategy) resulted in the formation of hydrogels with a uniform distribution of the functional molecules. Postmodification of the functional molecules of the C-SA-SC hydrogel with biotin or SpyTag could alter the three-dimensional (3D) spatial distribution of the functional molecules within the hydrogels depending on the mode of conjugation (SA/biotin or SpyCatcher/SpyTag), the size of the functional molecules, and the length of time of the modification. NIH-3T3 cells cultured on a C-SA-SC hydrogel, dual-functionalized with a biotinylated-Arg-Gly-Asp-Ser (RGDS) peptide and a basic fibroblast growth factor (bFGF) with SpyTag, showed cell adhesion to the PEG-SH-based hydrogels and cell morphological changes in response to the immobilized RGDS peptide and the bFGF. Moreover, the cells showed higher proliferation on the dual-functionalized C-SA-SC hydrogel than the cells cultured on hydrogels without either the RGDS peptide or the bFGF, demonstrating the benefits of dual-functionalizable hydrogels. The C-SA-SC hydrogel presented in this study is capable of being orthogonally functionalized by two different functional molecules with different 3D distributions of each molecule within the hydrogel and thus has the potential for use as a cell culturing scaffold for creating artificial cellular microstructures.
AB - Hydrogels possessing the ability to control cell functions have great potential as artificial substrates for cell culture. Herein, we report dual-functionalizable protein-polymer hybrid hydrogels prepared by thiol oxidation catalyzed by horseradish peroxidase and a phenolic molecule. A chimera protein of streptavidin (SA) and the SpyCatcher protein, with a cysteine residue at its N-terminus, (C-SA-SC) was constructed and co-cross-linked with thiol-functionalized four-arm polyethylene glycol (PEG-SH) to obtain hydrogels possessing two orthogonal conjugation moieties. Hydrogel formation using C-SA-SC conjugated with biotinylated or SpyTagged functional molecules (premodification strategy) resulted in the formation of hydrogels with a uniform distribution of the functional molecules. Postmodification of the functional molecules of the C-SA-SC hydrogel with biotin or SpyTag could alter the three-dimensional (3D) spatial distribution of the functional molecules within the hydrogels depending on the mode of conjugation (SA/biotin or SpyCatcher/SpyTag), the size of the functional molecules, and the length of time of the modification. NIH-3T3 cells cultured on a C-SA-SC hydrogel, dual-functionalized with a biotinylated-Arg-Gly-Asp-Ser (RGDS) peptide and a basic fibroblast growth factor (bFGF) with SpyTag, showed cell adhesion to the PEG-SH-based hydrogels and cell morphological changes in response to the immobilized RGDS peptide and the bFGF. Moreover, the cells showed higher proliferation on the dual-functionalized C-SA-SC hydrogel than the cells cultured on hydrogels without either the RGDS peptide or the bFGF, demonstrating the benefits of dual-functionalizable hydrogels. The C-SA-SC hydrogel presented in this study is capable of being orthogonally functionalized by two different functional molecules with different 3D distributions of each molecule within the hydrogel and thus has the potential for use as a cell culturing scaffold for creating artificial cellular microstructures.
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U2 - 10.1021/acsabm.0c00940
DO - 10.1021/acsabm.0c00940
M3 - Article
AN - SCOPUS:85096023964
SN - 2576-6422
VL - 3
SP - 7734
EP - 7742
JO - ACS Applied Bio Materials
JF - ACS Applied Bio Materials
IS - 11
ER -