title: Non-chromatographic purification of recombinant elastin-like polypeptides and their fusions with peptides and proteins from Escherichia coli.
abstract: Elastin-like polypeptides are repetitive biopolymers that exhibit a lower critical solution temperature phase transition behavior, existing as soluble unimers below a characteristic transition temperature and aggregating into micron-scale coacervates above their transition temperature. The design of elastin-like polypeptides at the genetic level permits precise control of their sequence and length, which dictates their thermal properties. Elastin-like polypeptides are used in a variety of applications including biosensing, tissue engineering, and drug delivery, where the transition temperature and biopolymer architecture of the ELP can be tuned for the specific application of interest. Furthermore, the lower critical solution temperature phase transition behavior of elastin-like polypeptides allows their purification by their thermal response, such that their selective coacervation and resolubilization allows the removal of both soluble and insoluble contaminants following expression in Escherichia coli. This approach can be used for the purification of elastin-like polypeptides alone or as a purification tool for peptide or protein fusions where recombinant peptides or proteins genetically appended to elastin-like polypeptide tags can be purified without chromatography. This protocol describes the purification of elastin-like polypeptides and their peptide or protein fusions and discusses basic characterization techniques to assess the thermal behavior of pure elastin-like polypeptide products.
authors: Department of Biomedical Engineering, Duke University; Research Triangle MRSEC, Duke University.
authors: Sarah R
authors: SR
authors: MacEwan
authors: Department of Biomedical Engineering, Duke University; Research Triangle MRSEC, Duke University.
authors: Wafa
authors: W
authors: Hassouneh
authors: Department of Biomedical Engineering, Duke University; Research Triangle MRSEC, Duke University; chilkoti@duke.edu.
authors: Ashutosh
authors: A
authors: Chilkoti
mesh_terms: D004549:Elastin / Q000737:chemistry / Q000302:isolation & purification*; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000302:isolation & purification*; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D010455:Peptides; D011506:Proteins; D011993:Recombinant Fusion Proteins; D004549:Elastin