Event Abstract Back to Event Enhancing the biological functions of non-animal collagens Yong Peng1*, Violet Stoichevska1*, Aditya V. Vashi1*, Jerome A. Werkmeister1*, Geoff J. Dumsday1* and John A. Ramshaw1* 1 CSIRO, CSIRO Manufacturing, Australia Introduction: Collagen-based biomedical materials have developed into important, clinically effective materials used in a range of devices that have gained wide acceptance. Recent developments have included the production of recombinant collagen materials that are well defined and are disease free. Most recently, a group of bacterial, non-animal collagens has emerged that may provide an excellent, safe and novel source of collagen for use in biomedical materials. These proteins form stable triple-helical structures without the need for secondary modification to include hydroxyproline in the sequence. However, these non-animal collagens are limited as they lack the binding motifs that are present in animal collagens and changes that require secondary modifications are generally not easily achieved. We have introduced multiple binding sites through sequence modifications that encompass up to 4 collagen triplets and also via sequences modified to include Cys and Tyr residue(s). Materials and Methods: Constructs were based on bacterial collagen structures obtained from GenBank records. Sequences encoding the globular and collagen-like sequences but lacking the C-terminal attachment domain were used. A His6-tag was included to assist purification. DNA samples were synthesised commercially. Sequences were modified to include mammalian heparin and integrin binding sequences and also to include GKY, GYC, GCC, GCP or GCP sequences. Expression was in E. coli using pCold vectors. Protein purification used Ni loaded HyperCel-Sepharose affinity chromatography with imidazole elution and further purification on a Sephacryl S200 26/60 column. Chemical modification reactions followed standard protocols. Results and Discussion: The non-animal collagens are effectively expressed in E. coli in high yield and were readily purified. Sequences that had been modified to include mammalian domains all exhibited the expected functions. Inclusion of unusual amino acids, cyclic peptides, sugars, lipids and other complex functions was achieved chemically through modifications of introduced Tyr and Cys residues. These introduced residues, which are not otherwise present, allowed a range of specific chemical modification reactions at defined locations. This approach was illustrated by a range of model reactions. These modifications include the use of Tyr additions and substitutions to allow gel formation, and Cys additions and substitutions to allow bromoacetyl, maleimidyl and vinyl sulfone based modifications. These point modifications enabled inclusion of alkyne (or azide) functions that allow the extensive range of substitutions that are available via ‘click’ chemistry to be accessed. Alternatively, when bifunctional reagents were used, this led to crosslinking to give higher molecular weight polymeric proteins. Higher molecular aggregates are useful in fabrication of sponges, as the non-animal collagens studied to date are typically smaller than animal interstitial collagens, around one quarter the length, and do not pack into large fibrillar aggregates like those that are formed by the major animal interstitial collagens. The higher molecular weight aggregates allow for more mechanically and dimensionally stable sponges. Conclusions: These data show that the functional limitations that may be present for certain applications of non-animal collagens are readily overcome by changes to the sequences of the recombinant proteins and the use, when needed, of specific chemical modifications at selected locations. Keywords: Tissue Engineering, protein, material design, biomacromolecule Conference: 10th World Biomaterials Congress, Montréal, Canada, 17 May - 22 May, 2016. Presentation Type: Poster Topic: Synthetic scaffolds as extracellular matrices Citation: Peng Y, Stoichevska V, Vashi AV, Werkmeister JA, Dumsday GJ and Ramshaw JA (2016). Enhancing the biological functions of non-animal collagens. Front. Bioeng. Biotechnol. Conference Abstract: 10th World Biomaterials Congress. doi: 10.3389/conf.FBIOE.2016.01.01158 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. 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Received: 27 Mar 2016; Published Online: 30 Mar 2016. * Correspondence: Dr. Yong Peng, CSIRO, CSIRO Manufacturing, Clayton, Australia, Email1 Dr. Violet Stoichevska, CSIRO, CSIRO Manufacturing, Clayton, Australia, violet.stoichevska@csiro.au Dr. Aditya V Vashi, CSIRO, CSIRO Manufacturing, Clayton, Australia, aditya.vashi@csiro.au Dr. Jerome A Werkmeister, CSIRO, CSIRO Manufacturing, Clayton, Australia, jerome.werkmeister@csiro.au Dr. Geoff J Dumsday, CSIRO, CSIRO Manufacturing, Clayton, Australia, geoff.dumsday@csiro.au Dr. John A Ramshaw, CSIRO, CSIRO Manufacturing, Clayton, Australia, john.ramshaw@csiro.au 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 Yong Peng Violet Stoichevska Aditya V Vashi Jerome A Werkmeister Geoff J Dumsday John A Ramshaw Google Yong Peng Violet Stoichevska Aditya V Vashi Jerome A Werkmeister Geoff J Dumsday John A Ramshaw Google Scholar Yong Peng Violet Stoichevska Aditya V Vashi Jerome A Werkmeister Geoff J Dumsday John A Ramshaw PubMed Yong Peng Violet Stoichevska Aditya V Vashi Jerome A Werkmeister Geoff J Dumsday John A Ramshaw 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.