Proteins secreted from cells are efficiently bound to and released from ECMs during injured tissue regeneration if the ECMs remain intact after injury. For example, the peptide domains of placenta growth factor-2 and laminin α subunit have high affinity to ECMs and induce signal transduction from their receptors, enhancing tissue repair. However, after severe tissue injury, a secreted protein needs to be provided along with the ECMs. Here, incorporating and releasing secreted proteins are generally incompatible, and thus the development of peptide-based materials capable of incorporating and releasing secreted proteins has been challenging. Cell-adhesive fibre-forming peptides with jigsaw-shaped hydrophobic surfaces (JigSAP) 18 can be used for efficient incorporation and sustained release of proteins. JigSAP incorporating JigSAP-tagged vascular endothelial growth factor (VEGF) shows regenerative therapeutic effects in stroke models without cell transplantation. AXXXA and GXXXG amino acid sequences are structural motifs found in glycophorin A. These motifs show α-helix-to-β-strand conformational transitions, generating a jigsaw-shaped hydrophobic surface that dimerizes with dovetail packing. This characteristic hydrophobic dimerization prompts nanofibre formation through β-sheet assembly. JigSAP, consisting of Ac-RIDARMRADIR-NH2 sequence, possesses the AXXXA motif in the hydrophobic part and shows one-dimensional self-assembly in aqueous media in silico calculated by MD simulation (Fig. ). When JigSAP (1 wt%) is dispersed in Dulbecco's modified Eagle's medium (DMEM), circular dichroism (CD) spectroscopic analysis visualizes a conformational change from helical or coil forms to β-sheet over 100 hours. In the transition to β-sheet, JigSAP forms nanofibres with an average width of 3.2 nm and length of several µm (Fig. ), and the dispersion becomes a hydrogel (Fig. ). Interestingly, the JigSAP hydrogel exhibits a remarkably high rigidity (Fig. 1.1 x 10 4 Pa), and the storage modulus G' and loss modulus G" of the JigSAP hydrogel monotonically decrease with increasing shear strain in the rheological analysis (Fig. ). The monotonic decrease of G" results in a strain thinning profile that is typically observed in covalently polymerized materials; therefore, this shows that supramolecular nanofibres of JigSAP are robust. When full-length enhanced green fluorescent protein (EGFP) is covalently conjugated with JigSAP (EGFP-JigSAP), 5V (EGFP-5V), and RADA16 (EGFP-RADA16) at the C-terminus (Fig. ), there is a dramatic difference in incorporation efficiency between JigSAP-tagged and non-tagged EGFP proteins; EGFP-JigSAP is incorporated at 93 mol%, whereas non-tagged EGFP is incorporated at only 3 mol% (Fig. ). Similarly, EGFP proteins tagged with 5V and RADA16 show higher incorporation efficiency into the corresponding peptide nanofibres than untagged EGFP (Fig. and, EGFP-5V: 66 mol%, EGFP-RADA16: 55 mol%). Importantly, the JigSAP hydrogel shows more efficient long-term release of the incorporated peptide-tagged proteins than the hydrogels consisting of other self-assembling peptides. The protein release profile from the gel shows that 21 mol% of EGFP-JigSAP incorporated into the JigSAP nanofibres is released after 24-hour incubation at 37 °C, and the percentage of released protein increases to 39 mol% after 168 hours of incubation (Fig. ). In contrast, the release of EGFP-5V and EGFP-RADA16 stagnates after 48 hours of incubation, and only 5 mol% and 8 mol%, respectively, are released even after 168 hours (Fig. ).