Fig 1.
SBA-NiO, a mesoporous silica material decorated with NiO nanoparticles, as the scaffolding support for protein fragments to interact.
(A) Schematic illustration of the interaction of protein half fragments induced by site-specific interaction of His-tags with NiO nanoparticles in the cavities of mesoporous SBA material. (B) TEM image of as synthesized SBA-15. (C) TEM image of SBA-NiO.
Fig 2.
(A) Schematic illustration of protein-fragment complementary assay (PCA) using mCherry as a “reporter” protein. (B) Split expression of mCherry at 159–160 position and at 136–137 position give two pairs of fragments N1/C1 and N2/C2 respectively (the structure of mCherry is drawn based on PDB ID 2H5Q) [37]. (C) SDS-PAGE picture of N1-mCherry and C1-mCherry. (D) Normalized photoluminescence (PL) spectra of mCherry (a), N1-mCherry (b), C1-mCherry (c), N1-mCherry + C1-mCherry (d) in solution at 1 mg/ml (λex = 587 nm).
Fig 3.
mCherry fragments assembled on SBA-NiO to form fluorescent protein.
(A) Schematic illustration of the assembly of mCherry fragments on the surface of SBA-NiO. (B) Fluorescent microscope image of N1-mCherry/C1-mCherry/SBA-NiO (left, TRITC channel; right, bright field). (C) Normalized fluorescent spectra of N1-mCherry/C1-mCherry/SBA-NiO (a), N1-mCherry/SBA-NiO (b), and N1-mCherry/N2-mCherry/SBA-NiO (c) (λex = 587 nm). All other assemblies do not fluorescence. (D) Quantification of PL intensity per μm2. a, SBA-NiO; b, BSA/SBA-NiO; c, N2-mCherry/C2-mCherry/SBA-NiO; d, N1-mCherry/N2-mCherry/SBA-NiO; e, N1-mCherry/SBA-NiO; f, N1-mCherry/C1-mCherry/SBA-NiO. (E) The PL intensity per area of N1-mCherry/C1-mCherry/SBA-NiO correlated with the amount of immobilized N1-mCherry and C1-mCherry.
Fig 4.
Split fragments of luciferase.
(A) Luciferase assay. (B) Split luciferase to give two complementary fragments N-Luc 2–416 and C-Luc 398–550, each his-tagged at N terminus and C terminus respectively (the structure is from PDB ID 2D1S) [39]. (C) SDS-PAGE picture of N-Luc and C-Luc.
Fig 5.
Fragments of Luciferase assembled on SBA-NiO to form functional luciferase.
(A) The assembly N-Luc/C-Luc/SBA-NiO catalyzed luciferin conversion, whereas N-Luc + C-Luc in solution remained inactive. a, N-Luc + C-Luc in solution; b, N-Luc/C-Luc/SBA-NiO. (B) Luciferase fragments alone on SBA-NiO N-Luc/SBA-NiO and C-Luc/SBA-NiO can regenerate the enzymatic activity. a, SBA-NiO; b, BSA/SBA-NiO; c, N-Luc/SBA-NiO; d, C-Luc/SBA-NiO; e, N-Luc/C-Luc/SBA-NiO. (C) Luciferase activity of N-Luc/C-Luc/SBA-NiO correlated with the amount of immobilized N-Luc and C-Luc.