Gene cassette design, cloning and expression of recombinant elastin like polypeptide to produce a functional biomaterial in tissue engineering

authors:

avatar Mohammad Reza Soleyman , avatar Mostafa khalili , avatar Alireza Soleyman Meigoni , avatar Hadi Mohammadzadeh Ghasghaii , avatar Adib Zendedel , avatar Maryam Baazm ORCID , *

Koomesh: Vol.17, issue 4; 1006-1016
article type: issue_documents_importer
received: August 01, 2015
accepted: April 01, 2016

how to cite: Soleyman M R, khalili M, Soleyman Meigoni A, Mohammadzadeh Ghasghaii H, Zendedel A, et al. Gene cassette design, cloning and expression of recombinant elastin like polypeptide to produce a functional biomaterial in tissue engineering. koomesh. 2024;17(4):e151196. 

Abstract

Introduction: Elastin-like polypeptide (ELP) s an artificial biopolymer and a pentapeptide with repeating motif of Val-Pro-Gly-Xaa-Gly (VPGXG). The structure is derived from extracellular cellular matrix (ECM) elastin. Protein-based polymers, which are composed of repeat units of natural or unnatural amino acids, have recently emerged as a promising new class of materials. The unique properties of ELPs such as being genetically encoded in a heterologous host (e.g., bacteria or eukaryotic cell), excitability under the influence of environmental factors, biocompatibility and biodegradability made them popular in a wide variety of biomedical applications. Materials and Methods: In this study, 330bp GRGDS-ELP monomer gene (expressing21 pentapeptide) was oligomerized into the cloning vector pUC to produce 990bp trimer gene (expressing63 fusion pentapeptide).After confirming gene structure by sequencing, the gene sequence was subcloned into pET-MOD expression vector and transduced into the E.coli. Results: Recombinant protein expression was induced by IPTG and the accuracy of the recombinant protein was confirmed by SDS-PAGE and Western blotting. Conclusion: With mentioned method, ELP was obtained for further studies

References

  • 1.

    Chow D, Nunalee ML, Lim DW, Simnick AJ, Chilkoti A. Peptide-based biopolymers in biomedicine and biotechnology. Mater Sci Eng R Rep 2008; 62: 125-155.

  • 2.

    O'Brien FJ. Biomaterials & scaffolds for tissue engineering. Materials Today 2011; 14: 88-95.

  • 3.

    Bandiera A1, Sist P, Urbani R. Comparison of thermal behavior of two recombinantly expressed human elastin-like polypeptides for cell culture applications. Biomacromolecules 2010; 11: 3256-3265.

  • 4.

    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; 3: 357-367.

  • 5.

    Chilkoti A, Dreher MR, Meyer DE. Design of thermally responsive, recombinant polypeptide carriers for targeted drug delivery. Adv Drug Deliv Rev 2002; 54: 1093-1111.

  • 6.

    Mithieux SM, Weiss AS. Elastin. Adv Protein Chem 2005; 70: 437-461.

  • 7.

    Kagan HM, Sullivan KA. Lysyl oxidase: preparation and role in elastin biosynthesis. Methods Enzymol 1981; 82: 637-650.

  • 8.

    Liu JM, Davidson JM. The elastogenic effect of recombinant transforming growth factor-beta on porcine aortic smooth muscle cells. Biochem Biophys Res Commun 1988; 154: 895-901.

  • 9.

    Debelle L, Tamburro AM. Elastin: molecular description and function. Int J Biochem Cell Biol 1999; 31: 261-272.

  • 10.

    Floss DM, Schallau K, Rose-John S, Conrad U, Scheller J. Elastin-like polypeptides revolutionize recombinant protein expression and their biomedical application. Trends Biotechnol 2010; 28: 37-45.

  • 11.

    Meyer DE, Chilkoti A. Purification of recombinant proteins by fusion with thermally-responsive polypeptides. Nature Biotechnol 1999; 17: 1112-1115.

  • 12.

    MacEwan SR, Chilkoti A. Applications of elastin-like polypeptides in drug delivery. J Control Release 2014; 190: 314-330.

  • 13.

    Saxena R, Nanjan MJ. Elastin-like polypeptides and their applications in anticancer drug delivery systems: a review. Drug Deliv 2015; 22: 156-167.

  • 14.

    Kyle S, Aggeli A, Ingham E, McPherson MJ. Production of self-assembling biomaterials for tissue engineering. Trends Biotechnol 2009; 27: 423-433.

  • 15.

    Pierschbacher MD, Ruoslahti E. Cell attachment activity of fibronectin can be duplicated by small synthetic fragments of the molecule. Nature 1983; 309: 30-33.

  • 16.

    Christensen T, Trabbic-Carlson K, Liu W, Chilkoti A. Purification of recombinant proteins from E. coli at low expression levels by inverse transition cycling. Anal Biochem 2007; 360: 166-168.

  • 17.

    Hwang DS, Sim SB, Cha HJ. Cell adhesion biomaterial based on mussel adhesive protein fused with RGD peptide. Biomaterials 2007; 28: 4039-4046.

  • 18.

    Hersel U, Dahmen C, Kessler H. RGD modified polymers: biomaterials for stimulated cell adhesion and beyond. Biomaterials 2003; 24: 4385-4415.

  • 19.

    Pierschbacher MD, Ruoslahti E. Influence of stereochemistry of the sequence Arg-Gly-Asp-Xaa on binding specificity in cell adhesion. J Biol Chem 1987; 262: 17294-17298.

  • 20.

    Blindt R, Vogt F, Astafieva I, Fach C, Hristov M, Krott N, et al. A novel drug-eluting stent coated with an integrin-binding cyclic Arg-Gly-Asp peptide inhibits neointimal hyperplasia by recruiting endothelial progenitor cells. J Am Coll Cardiol 2006; 47: 1786-1795.

  • 21.

    Ferreras M, Gavilanes JG, Garca-Segura JM. A permanent Zn< sup> 2+ reverse staining method for the detection and quantification of proteins in polyacrylamide gels. Anal Biochem 1993; 213: 206-212.

  • 22.

    Abdelaal OA, Darwish SM. Fabrication of tissue engineering scaffolds using rapid prototyping techniques. WASET 2011; 59: 577-585.

  • 23.

    Silva R, Fabry B, Boccaccini AR. Fibrous protein-based hydrogels for cell encapsulation. Biomaterials 2014; 35: 6727-6738.

  • 24.

    McDaniel JR, Callahan DJ, Chilkoti A. Drug delivery to solid tumors by elastin-like polypeptides. Adv Drug Deliv Rev 2010; 62: 1456-1467.

  • 25.

    Abdelaal OA, Darwish SM. Fabrication of tissue engineering scaffolds using rapid prototyping techniques. World Academy of Science, Engineering and Technology, Int J Mechanical, Aerospace, Industrial, Mechatronic and Manufacturing Engineering 2011; 5: 2317-2325.

  • 26.

    Nettles DL, Chilkoti A, Setton LA. Applications of elastin-like polypeptides in tissue engineering. Adv Drug Deliv Rev 2010; 62: 1479-1485.

  • 27.

    Simnick AJ, et al. Biomedical and biotechnological applications of elastin-like polypeptides. Journal of Macromolecular Science, Part C: Polymer Reviews 2007; 47: 121-154.

  • 28.

    Altunbas A, Pochan DJ. Peptide-based and polypeptide-based hydrogels for drug delivery and tissue engineering. Top Curr Chem 2012; 310: 135-167.

  • 29.

    Girotti A, Reguera J, Rodrguez-Cabello JC, Arias FJ, Alonso M, Matestera A. Design and bioproduction of a recombinant multi (bio) functional elastin-like protein polymer containing cell adhesion sequences for tissue engineering purposes. J Mater Sci Mater Med 2004; 15: 479-484.

  • 30.

    Koria P, Yagi H, Kitagawa Y, Megeed Z, Nahmias Y, Sheridan R, Yarmush ML. Self-assembling elastin-like peptides growth factor chimeric nanoparticles for the treatment of chronic wounds. Proc Natl Acad Sci U S A 2011; 108: 1034-1039.

  • 31.

    Minato A, Ise H, Goto M, Akaike T. Cardiac differentiation of embryonic stem cells by substrate immobilization of insulin-like growth factor binding protein 4 with elastin-like polypeptides. Biomaterials 2012; 33: 515-523.

  • 32.

    Amiram M, Luginbuhl KM, Li X, Feinglos MN, Chilkoti A. Injectable protease-operated depots of glucagon-like peptide-1 provide extended and tunable glucose control. Proc Natl Acad Sci U S A 2013; 110: 2792-2797.

  • 33.

    Ryu JS, Raucher D. Anti-tumor efficacy of a therapeutic peptide based on thermo-responsive elastin-like polypeptide in combination with gemcitabine. Cancer lett 2014; 348: 177-184.

  • 34.

    Massodi I, Thomas E, Raucher D. Application of thermally responsive elastin-like polypeptide fused to a lactoferrin-derived peptide for treatment of pancreatic cancer. Molecules 2009; 14: 1999-2015.

  • 35.

    Lin HT, Chiou SH, Kao CL, Shyr YM, Hsu CJ, Tarng YW, et al. Characterization of pancreatic stem cells derived from adult human pancreas ducts by fluorescence activated cell sorting. World J Gastroenterol 2006; 12: 4529-4535.

  • 36.

    Takagi J. Structural basis for ligand recognition by RGD (Arg-Gly-Asp)-dependent integrins. Biochem Soc Trans 2004; 32: 403-406.

  • 37.

    Chen CS, Alonso JL, Ostuni E, Whitesides GM, Ingber DE. Cell shape provides global control of focal adhesion assembly. Biochem Biophys Res Commun 2003; 307: 355-361.

  • 38.

    Nicol A, Channe Gowda D, Urry DW. Cell adhesion and growth on synthetic elastomeric matrices containing ARGGLYASPSER3. J Biomed Material Res 1992; 26: 393-413.

  • 39.

    Liu JC, Tirrell DA. Cell response to RGD density in cross-linked artificial extracellular matrix protein films. Biomacromolecules 2008; 9: 2984-2988.

  • 40.

    Huang K, Duan N, Zou W, Zhang C, Lai Y, Shen P, Hua Z. Fused hydrophobic elastin-like-peptides (ELP) enhance biological activity of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Protein Pept Lett 2015; 22: 1000-1006.

  • 41.

    MacEwan SR, Hassouneh W, Chilkoti A. Non-chromatographic Purification of Recombinant Elastin-like Polypeptides and their Fusions with Peptides and Proteins from Escherichia coli. J Vis Exp 2014; 88.

  • 42.

    McPherson DT, Xu J, Urry DW. Product purification by reversible phase transition following escherichia coli expression of genes encoding up to 251 repeats of the elastomeric pentapeptide GVGVP. Protein Expr Purif 1996; 7: 51-57.