title: Recombinant production of self-assembling β-structured peptides using SUMO as a fusion partner.
abstract: BACKGROUND
  Self-assembling peptides that form nanostructured hydrogels are important biomaterials for tissue engineering scaffolds. The P₁₁-family of peptides includes, P₁₁-4 (QQRFEWEFEQQ) and the complementary peptides P₁₁-13 (EQEFEWEFEQE) and P₁₁-14 (QQOrnFOrnWOrnFOrnQQ). These form self-supporting hydrogels under physiological conditions (pH 7.4, 140 mM NaCl) either alone (P₁₁-4) or when mixed (P₁₁-13 and P₁₁-14). We report a SUMO-peptide expression strategy suitable for allowing release of native sequence peptide by SUMO protease cleavage.
  
  
  RESULTS
  We have expressed SUMO-peptide fusion proteins from pET vectors by using autoinduction methods. Immobilised metal affinity chromatography was used to purify the fusion protein, followed by SUMO protease cleavage in water to release the peptides, which were recovered by reverse phase HPLC. The peptide samples were analysed by electrospray mass spectrometry and self-assembly was followed by circular dichroism and transmission electron microscopy.
  
  
  CONCLUSIONS
  The fusion proteins were produced in high yields and the β-structured peptides were efficiently released by SUMO protease resulting in peptides with no additional amino acid residues and with recoveries of 46% to 99%. The peptides behaved essentially the same as chemically synthesised and previously characterised recombinant peptides in self-assembly and biophysical assays.
authors: Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, Leeds LS2 9JT, UK.
authors: Abhinav
authors: A
authors: Prakash
authors: Stephen J
authors: SJ
authors: Parsons
authors: Stuart
authors: S
authors: Kyle
authors: Michael J
authors: MJ
authors: McPherson
mesh_terms: D000595:Amino Acid Sequence; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D056148:Chromatography, Reverse-Phase; D004926:Escherichia coli / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate / Q000737:chemistry; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D025841:Small Ubiquitin-Related Modifier Proteins / Q000235:genetics / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D010455:Peptides; D011993:Recombinant Fusion Proteins; D025841:Small Ubiquitin-Related Modifier Proteins; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate