Novel nanocomposites from spider silk–silica fusion (chimeric) proteins
- Cheryl Wong Po Foo*,
- Siddharth V. Patwardhan†,
- David J. Belton†,
- Brandon Kitchel*,
- Daphne Anastasiades*,
- Jia Huang*,
- Rajesh R. Naik‡,
- Carole C. Perry†,§, and
- David L. Kaplan*,¶
- *Departments of Biomedical Engineering, Chemistry, and Chemical and Biological Engineering, Bioengineering and Biotechnology Center, Tufts University, Medford, MA 02155;
- †Biomolecular and Materials Interface Research Group, School of Biomedical and Natural Sciences, Nottingham Trent University, Nottingham NG11 8NS, United Kingdom; and
- ‡Materials and Manufacturing Directorate, Air Force Research Laboratory, 3005 Hobson Way, Wright–Patterson Air Force Base, OH 45433-7702
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Edited by Charles R. Cantor, Sequenom, Inc., San Diego, CA, and approved May 8, 2006 (received for review February 10, 2006)
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Fig. 1.Schematic representation of the design of fusion proteins and their use in controlled silica nanocomposite formation. (A) Scheme of chimeric design with two functional domains: silk and R5. (B) Model of spider silk protein processing into films and fibers and silicification reactions on the assembled materials.
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Fig. 2.Expression and purification of spider silk fusion proteins. Amino acid sequences and gel electrophoresis of the bioengineered spider silk fusion proteins CRGD15mer+R5 (A) and 15mer+R5 (B). Underlined is the representative monomeric repeat unit selected and used in the design of the recombinant proteins based on the consensus sequence of spidroin 1 (Masp1) native sequence of N. clavipes (GenBank accession no. P19837). (C) Gel electrophoresis of 15mer+R5: lane 1, ladder; lane 2, flow-through; lane 3, wash 1; lane 4, elution 1; lane 5, elution 2. (D) Gel electrophoresis of CRGD15mer+R5: lanes 1, ladder; lane 2, elution 1; lane 3, elution 2; lane 4, elution 3 treated with 10 mM DTT.
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Fig. 3.Effect of R5 peptide and fusion proteins on silicification. (A) Dynamic light scattering data for silica obtained from experiments performed in the presence of R5 alone. (B) Typical SEM of silica formed in the presence of 1 silicon:1 nitrogen-containing amino acid in R5 at pH 7.0 from aqueous solution. The ratio of silicon to nitrogen containing amino acid side chains in R5 was kept constant at 1:1 to allow comparison with our previous studies using simple amino acids, peptides, and small amines (44, 45). (C) Normalized absorbance from soluble silicon released from a 30 mM aqueous solution of a silicon catecholato complex in the presence and absence of fusion proteins (silk+R5) over 24 h at pH adjusted to ≈6.8 as described previously (44, 45).
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Fig. 4.Morphological and elemental analyses of nanocomposites. SEM images of silica composite materials generated from an aqueous-based dipotassium silicon triscatecholate complex in the presence of chimera CRGD15mer+R5 (A and B) and chimera 15mer+R5 (C and D). Areas highlighted by rectangles are presented at higher magnification in the same rows. In A–C elemental maps are shown for silicon (Si), oxygen (O), and/or carbon (C).
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Fig. 5.SEM images of untreated and methanol-treated silk films formed from four different genetically engineered silk proteins: CRGD15mer, 15mer, CRGD15mer+R5, and 15mer+R5. Images of the control films and the films that underwent silicification reactions are shown.
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Fig. 6.Morphological characterization and elemental analysis of silica deposition on eletrospun fibers. (A) SEM images of untreated and treated electrospun CRGD15mer+R5 silk fibers before, during, and after silicification reactions. (B and C) X-ray photoelectron spectroscopy analysis of CRGD15mer+R5 and silicified CRGD15mer+R5 on Al foil and on silicon chip at the characteristic binding energies of 153 eV (B) and 102 eV (C) for electrons found in the 2s and 2p3 electron shells of the silicon atom, respectively.
Footnotes
- §To whom correspondence may be addressed at: Interdisciplinary Biomedical Research Centre, School of Biomedical and Natural Sciences, Nottingham Trent University, Clifton Lane, Nottingham NG11 8NS, United Kingdom . E-mail: carole.perry{at}ntu.ac.uk
- ¶To whom correspondence may be addressed at: Department of Biomedical Engineering, Bioengineering and Biotechnology Center, Tufts University, 4 Colby Street, Medford, MA 02155 . E-mail: david.kaplan{at}tufts.edu
- © 2006 by The National Academy of Sciences of the USA
