Expression, cross-linking and characterization of recombinant chitin binding resilin

Guokui Qin, Shaul Lapidot, Keiji Numata, Xiao Hu, Sigal Meirovitch, Mara Dekel, Itai Podoler, Oded Shoseyov, David L. Kaplan

Research output: Chapter in Book/Report/Conference proceedingConference contribution

Abstract

Resilin is a polymeric rubber-like protein secreted by insects to specialized cuticle regions, in areas where high resilience and low stiffness are required. Resilin binds to the cuticle polysaccharide chitin via a chitin binding domain and is further polymerized through oxidation of the tyrosine residues resulting in the formation of dityrosine bridges and assembly of a high-performance protein-carbohydrate composite material. We describe for the first time a comprehensive study on the mechanical, structural and biochemical function of chitin binding recombinant Drosophila melanogaster resilin. Various resilin constructs were cloned including the full length gene enabling Ni-NTA purification, as well as heat and salt precipitation for rapid and efficient purification. The binding isotherms and constants (Kd, B max) of resilin to chitin via its chitin binding domain were determined and displayed high affinity to chitin, implying its important role in the assembly of the resilin-chitin composite. The structural and elastic properties were investigated using Fourier Transform Infrared Spectroscopy (FTIR), Circular Dichroism (CD) and Atomic Force Microscopy (AFM) with peroxidase crosslinked solid resilin materials. Generally, little structural organization was found by these biophysical methods, suggesting structural order was not induced by the dityrosine crosslinks. Further, the elastomeric properties found from the full length protein compared favorably with the shorter resilin generated previously from exon 1. The unusual elastomeric behavior of this protein suggests possible utility in biomaterials applications.

Original languageEnglish (US)
Title of host publicationProceedings of the 2010 IEEE 36th Annual Northeast Bioengineering Conference, NEBEC 2010
DOIs
StatePublished - Jun 8 2010
Event36th Annual Northeast Bioengineering Conference, NEBEC 2010 - New York, NY, United States
Duration: Mar 26 2010Mar 28 2010

Publication series

NameProceedings of the 2010 IEEE 36th Annual Northeast Bioengineering Conference, NEBEC 2010

Other

Other36th Annual Northeast Bioengineering Conference, NEBEC 2010
CountryUnited States
CityNew York, NY
Period3/26/103/28/10

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All Science Journal Classification (ASJC) codes

  • Bioengineering

Cite this

Qin, G., Lapidot, S., Numata, K., Hu, X., Meirovitch, S., Dekel, M., Podoler, I., Shoseyov, O., & Kaplan, D. L. (2010). Expression, cross-linking and characterization of recombinant chitin binding resilin. In Proceedings of the 2010 IEEE 36th Annual Northeast Bioengineering Conference, NEBEC 2010 [5458181] (Proceedings of the 2010 IEEE 36th Annual Northeast Bioengineering Conference, NEBEC 2010). https://doi.org/10.1109/NEBC.2010.5458181