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J Am Chem Soc. 2015 Sep 9;137(35):11285-93. doi: 10.1021/jacs.5b03593. Epub 2015 Jul 14.

Self-Assembling Nano-Architectures Created from a Protein Nano-Building Block Using an Intermolecularly Folded Dimeric de Novo Protein.

Author information

1
Japan Society for the Promotion of Science , Chiyoda, Tokyo 102-8471, Japan.
2
Graduate School of Medical Life Science, Yokohama City University , Tsurumi, Yokohama, Kanagawa 230-0045, Japan.
3
Department of Chemistry, Princeton University , Princeton, New Jersey 08544, United States.
4
Institute for Biomedical Sciences, Interdisciplinary Cluster for Cutting Edge Research, Shinshu University , Matsumoto, Nagano 390-8621, Japan.

Abstract

The design of novel proteins that self-assemble into supramolecular complexes is an important step in the development of synthetic biology and nanotechnology. Recently, we described the three-dimensional structure of WA20, a de novo protein that forms an intermolecularly folded dimeric 4-helix bundle (PDB code 3VJF ). To harness the unusual intertwined structure of WA20 for the self-assembly of supramolecular nanostructures, we created a protein nanobuilding block (PN-Block), called WA20-foldon, by fusing the dimeric structure of WA20 to the trimeric foldon domain of fibritin from bacteriophage T4. The WA20-foldon fusion protein was expressed in the soluble fraction in Escherichia coli, purified, and shown to form several homooligomeric forms. The stable oligomeric forms were further purified and characterized by a range of biophysical techniques. Size exclusion chromatography, multiangle light scattering, analytical ultracentrifugation, and small-angle X-ray scattering (SAXS) analyses indicate that the small (S form), middle (M form), and large (L form) forms of the WA20-foldon oligomers exist as hexamer (6-mer), dodecamer (12-mer), and octadecamer (18-mer), respectively. These findings suggest that the oligomers in multiples of 6-mer are stably formed by fusing the interdigitated dimer of WA20 with the trimer of foldon domain. Pair-distance distribution functions obtained from the Fourier inversion of the SAXS data suggest that the S and M forms have barrel- and tetrahedron-like shapes, respectively. These results demonstrate that the de novo WA20-foldon is an effective building block for the creation of self-assembling artificial nanoarchitectures.

PMID:
26120734
DOI:
10.1021/jacs.5b03593
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