Elastin-like polypeptides
Elastin-like polypeptides (ELPs) are synthetic biopolymers with potential applications in the fields of cancer therapy, tissue scaffolding, metal recovery, and protein purification.
Specifically, the hydrophilicity or hydrophobicity and the presence or absence of a charge on the guest residue play a great role in determining the Tt.
The repeat sequences found in the biopolymer give each ELP a distinct structure, as well as influence the lower critical solution temperature (LCST), also referred to commonly as the Tt.
These hydrophobic domains, consisting overwhelmingly of alanine, proline, glycine, and valine, tend towards instability and disorderliness, ensuring that the elastin does not lock into any specific confirmation.
Thus, ELPs consisting of the Val-Pro-Gly-X-Gly monomeric units, which bear resemblance to the repetitive tropoelastin hydrophobic domains, are highly disordered below their Tt.
Glycine, due to the lack of a bulky side chain, enables the biopolymer to be flexible and proline prevents the formation of stable hydrogen bonds in the ELP backbone.
[7] The transition temperature of an ELP depends to a certain extent on the identity of the "X" residue found at the fourth position of the pentapeptide monomeric unit.
Summing up the contribution of each potential guest residue (XAA) will yield an SI index that is directly proportional to ΔTt,fusion.
It was found that the amino acids that are charged under a physiological pH of 7.4 have the greatest impact on the overall SI of a fused protein.
[10] Because ELPs are protein-based biopolymers, synthesis involves manipulation of genes to continually express the monomeric repeat unit.
[5][9] Also, ELPs can be experimentally modified through conjugation with other polymers or through SpyTag/SpyCatcher reaction,[11] allowing for the synthesis of copolymers with unique morphology.
[9] The OEPCR method uses a small amount of the gene encoding the monomeric ELP unit and leads to the amplification of this segment to a great extent.
[9] In recursive directional ligation, the gene encoding the monomer is inserted into a plasmid with restriction sites that are recognized by at least two endonucleases.
However, the restriction endonucleases used are limited to those that do not cut within the ELP monomer gene itself, as this would lead to loss of crucial nucleotides and a potential frameshift mutation in the protein.
Through a cycloaddition reaction involving both of the functional groups and manipulation of the solvent pH, diblock and star polymers can be formed.
To ensure that there are few impurities in the ELP-protein complex isolated, the solution can be cooled below the Tt, enabling the ELPs to once again assume their linear structure.
From this point, hot and cold centrifugation cycles can be repeated, and then the protein of interest can be eluted from the ELPs via the addition of a salt.
[3] The temperature-based phase behavior of ELPs can be utilized to produce stiff networks that may be compatible with cellular regeneration applications.
These otherwise brittle networks can then be modified chemically, via oxidative coupling, to yield hydrogels which can sustain high levels of mechanical stress and strain.
