Tropoelastin confers structural integrity, mechanical resilience, and biological properties to elastic tissues. Systematic mutational studies of tropoelastin reveal key regions critical for protein function. We couple this understanding of tropoelastin biology with established plasma treatment technology to stably impart biological activity to bio-inert metals and polymers commonly used for orthopaedic implants. These new bio-engineered materials elicit improved bone cell responses for bone repair and regeneration.
The tropoelastin molecule: Basic studies of protein structure and function
This part of the work explores the functional significance of specific regions in human tropoelastin. Mutant tropoelastin constructs were designed in which the R515 residue in the bridge region (1), the E345/E414 residues near the hinge region, or the D72 residue in the N-terminal region (2) have been inactivated by alanine substitution. Another mutant construct that contains domain 22, which is typically spliced out in the hinge region of native human tropoelastin, was also produced.
The tropoelastin mutants display varying degrees of impaired self-assembly. In collaboration with an international team of structural biologists and synchrotron scientists, the nanostructures of the mutant species were found to possess sub-molecular conformational changes consistent with their dysfunctional biochemical properties.
Until this time, tropoelastin has classically been considered a globally flexible protein, with pathological consequences often due to large deletions or insertions within the molecule. Our results, published in the Proceedings of the National Academy of Sciences and the Journal of Biological Chemistry, identify for the first time the role of specific, single residues in maintaining the shape of human tropoelastin, and the dramatic consequences of nano-scale structural changes to protein assembly and function.
Bringing together physics and molecular biology: Development of tropoelastin-functionalised materials
Armed with a more extensive understanding of tropoelastin biology, we wish to harness the bioactivity of this protein for medical applications (3). We can functionalise bioinert materials, including metals and polymers, by stably immobilising tropoelastin on their surface.
This technique relies on using plasma technology to chemically modify the material surface (4). Metals are modified by the deposition of a plasma-activated coating, which utilizes acetylene, nitrogen and argon plasma to form a cross-linked carbonised layer on the surface. Similarly, polymers are modified by bombardment with nitrogen plasma. These processes create reactive free radicals on the material surface that can stably bind biomolecules without the use of toxic chemical linkers. We have experimental validation that plasma-modified metals and polymers covalently bind tropoelastin in an active conformation.
From bench to clinic: Tropoelastin-functionalised materials for bone repair
Tropoelastin, as a major component of elastic tissue, has classically been used in soft tissue applications. Here, we explore its potential in a non-traditional context of bone repair.
Metals such as titanium and zirconium, and polymers such as polyetheretherketone (PEEK) are commonly used to manufacture bone implants because of their mechanical strength and biocompatibility. However, as bioinert materials, they do not actively promote bone integration and formation, which are integral to the long-term success of orthopaedic implants. We find that tropoelastin-functionalised zirconium and PEEK promote significantly higher bone cell adhesion, proliferation, matrix maturation, and mineral deposition compared to untreated materials. These processes are requisite to new bone formation. Our findings can be readily translated to the development of improved orthopedic implants for accelerated bone healing.
3. Additional Details
- Yeo, G. C., Baldock, C., Tuukkanen, A., Roessle, M., Dyksterhuis, L. B., Wise, S. G., Matthews, J., Mithieux, S. M., and Weiss, A. S. (2012) Tropoelastin bridge region positions the cell-interactive C terminus and contributes to elastic fiber assembly. Proceedings of the National Academy of Sciences 109, 2878-2883.
- Yeo, G. C., Baldock, C., Wise, S. G., and Weiss, A. S. (2014) A negatively charged residue stabilizes the tropoelastin N-terminal region for elastic fiber assembly. J Biol Chem 289, 34815-34826.
- Yeo, G. C., Aghaei-Ghareh-Bolagh, B., Brackenreg, E. P., Hiob, M. A., Lee, P., and Weiss, A. S. (2015) Fabricated Elastin. Adv Healthc Mater 13, 201400781.
- Wise, S. G., Waterhouse, A., Kondyurin, A., Bilek, M. M., and Weiss, A. S. (2012) Plasma-based biofunctionalization of vascular implants. Nanomedicine 7, 1907-1916.
- Wang, Y., Mithieux, S. M., Kong, Y., Wang, X. Q., Chong, C., Fathi, A., Dehghani, F., Panas, E., Kemnitzer, J., Daniels, R., Kimble, R. M., Maitz, P. K., Li, Z., and Weiss, A. S. (2015) Tropoelastin incorporation into a dermal regeneration template promotes wound angiogenesis. Adv Healthc Mater 4, 577-584.
- Hiob, M. A., Wise, S. G., Kondyurin, A., Waterhouse, A., Bilek, M. M., Ng, M. K., and Weiss, A. S. (2013) The use of plasma-activated covalent attachment of early domains of tropoelastin to enhance vascular compatibility of surfaces. Biomaterials 34, 7584-7591.