Developing self-assembling elastin-like polypeptide nanoparticles using medium-length sequences: ramifications of chain length and guest amino acid hydrophobicity on assembly and disassembly conditions

Event Abstract Back to Event Developing self-assembling elastin-like polypeptide nanoparticles using medium-length sequences: ramifications of chain length and guest amino acid hydrophobicity on assembly and disassembly conditions Markian Bahniuk1, 2, Abdullah K. Alshememry2, 3 and Larry D. Unsworth1, 2, 4 1 University of Alberta, Biomedical Engineering, Canada 2 Nationa Research Council, National Institute for Nanotechnology, Canada 3 University of Alberta, Pharmacy and Pharmaceutical Sciencecs, Canada 4 University of Alberta, Chemical and Materials Engineering, Canada Introduction: As medicine continues to enter the realm of the nanoscale, there is an increasing demand for smart, flexible and safe biomaterials for sophisticated applications. Due to their temperature-induced reversible self-assembly, nanoparticles formed from elastin-like polypeptides (ELPs) may be uniquely suited for drug delivery in conditions where temperature decreases are crucial, viz., hypothermic therapies[1]. ELPs are oligomers of the amino acid sequence VPGXG where X cannot be proline[2]. Most ELP work involves either peptides that can be manufactured using solid-phase synthesis (10 repeats or fewer) or large (160 repeats and above) proteins made using recombinant methods. The work herein describes the synthesis and characterization of medium length ELPs made up of 20-160 repeats and their behaviour as they self-assemble into nanoparticles. Materials and Methods: DNA sequences encoding ELP genes were assembled and the expression and purification followed standard protocols for polyhistidine-based affinity chromatography followed by inverse temperature cycling (ITC) to further purify the ELPs[3]. Solution properties were screened to produce particles, and it was found that in phosphate buffered saline at pH 7.4, ELPs formed particles at 0.4-2mg/mL depending on the specific construct. The particle assembly and disassembly were characterized using a Malvern Nano-ZS Zetasizer and a Hitachi S4800 high-resolution scanning electron microscope. DLS studies were performed duplicate by ramping the temperature by 5°C then equilibrating the sample for 2 minutes before measuring. Results: Conditions under which each ELP construct formed particles and disassociated varied (Figure 1). At temperatures increasing from 5 to 25°C, a leucine-containing ELP made of 20 repeats (L20) showed a particle diameter of approximately 5nm. This increased to about 600nm until the temperature reached 35°C at which point the particle size decreased to 450nm. Cooling this sample showed the same changes in size, but at lower temperatures than while heating. An L40 ELP exhibited only one increase in size at 15°C while heating and minimal dissolution behaviour upon cooling. A valine-containing ELP (V40) of similar length as L40 also showed only one increase in size with heating but at 25°C. This construct also showed dissolution behaviour at 20°C. Discussion: The choice of guest amino acid and/or number of repeats does play a role in the formation and dissolution properties of ELP nanoparticles. Decreasing chain lengths increase the temperature for assembly and dissolution, though perhaps not as drastically as hypothesized in the literature for this size range. More hydrophobic guest amino acid chemistries resulted in decreased transition temperatures by ~10°C as well as more noticeable hysteresis and particle stability upon cooling. The differences between the ELP constructs did not, however, result in any significant variance in size of the nanoparticles. Conclusion: The work herein describes the results of engineering ELPs to form nanoparticles by using constructs with atypical numbers of repeats. These ELPs do indeed form nanoparticles and variations in their sequences result in differences in their self-assembly and disassembly temperatures. This demonstrates that ELP-based nanoparticles are a robust, customizable biomaterial which can be engineered to behave in specific ways for a wide variety of biomaterial applications. The authors would like to acknowledge financial support provided by the National Research Council of Canada and National Institute for Nanotechnology, the University of Alberta and Alberta Innovates Technology Futures opportunity #200801440.; The authors would like to thank Lavina Ng, Catherine de Guzman and Dr. Jae-Young Cho for their various technical contributions during this study.References:[1] Urry DW. Physical Chemistry of Biological Free Energy Transduction As Demonstrated by Elastic Protein-Based Polymers. The Journal of Physical Chemistry B. 1997;101(51):11007-28[2] Urry DW, Trapane TL, Prasad KU. Phase-structure transitions of the elastin polypentapeptide-water system within the framework of composition-temperature studies. Biopolymers. 1985;24(12):2345-56[3] Meyer DE, Chilkoti A. Genetically Encoded Synthesis of Protein-Based Polymers with Precisely Specified Molecular Weight and Sequence by Recursive Directional Ligation: Examples from the Elastin-like Polypeptide System. Biomacromolecules. 2002:357-67 Keywords: self-assembly, nanoparticle, protein, Environmental response Conference: 10th World Biomaterials Congress, Montréal, Canada, 17 May - 22 May, 2016. Presentation Type: Poster Topic: Protein-based biomaterials Citation: Bahniuk M, Alshememry AK and Unsworth LD (2016). Developing self-assembling elastin-like polypeptide nanoparticles using medium-length sequences: ramifications of chain length and guest amino acid hydrophobicity on assembly and disassembly conditions. Front. Bioeng. Biotechnol. Conference Abstract: 10th World Biomaterials Congress. doi: 10.3389/conf.FBIOE.2016.01.00068 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 27 Mar 2016; Published Online: 30 Mar 2016. Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Markian Bahniuk Abdullah K Alshememry Larry D Unsworth Google Markian Bahniuk Abdullah K Alshememry Larry D Unsworth Google Scholar Markian Bahniuk Abdullah K Alshememry Larry D Unsworth PubMed Markian Bahniuk Abdullah K Alshememry Larry D Unsworth Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.