Production and Purification of a Recombinant Elastomeric Polypeptide, G‐(VPGVG)₁₉‐VPGV, from Escherichia coli

Abstract

An elastomeric polypeptide was produced, with the sequence G-(VPGVG)19-VPGV, as a fusion to glutathione S-transferase using the vector pGEX-3X. The fusion protein was expressed to high levels in Escherichia coli as indicated by SDS-PAGE analysis of induced cells. The fusion protein was affinity purified and cleaved with protease factor Xa, and the elastomeric polypeptide was recovered to a high degree of purity as indicated by SDS-PAGE followed by staining with CuCl2. The physical characterizations of carbon-13 and proton nuclear magnetic resonance and of the temperature profile for turbidity formation for the inverse temperature transition of hydrophobic folding and assembly attest to the successful microbial synthesis of the polypentapeptide of elastin. The results of these studies provide the initial progress toward achieving a more economical and practical means of producing material for elastic protein-based polymer research and applications.

Biocompatible ELR-Based Polyplexes Coated with MUC1 Specific Aptamers and Targeted for Breast Cancer Gene Therapy

“…This retard effect on electrophoretic mobility for ELRs when compared to a commercial MW marker in SDS-PAGE is well known and was previously described [1][2][3] This material is available free of charge via the Internet at http://pubs.acs.org.…”

Extensive neurite outgrowth and active synapse formation on self-assembling peptide scaffolds

“…Stable macroscopic matrix structures can be fabricated by the spontaneous self assembly of aqueous sapeptide solutions introduced to physiological salt-containing solutions. The self assembly of other sapeptide and protein materials has been described (9)(10)(11)(33)(34)(35)(36)(37)(38)(39). One peptide gel was reported by Aggeli et al, who found a fragment of a membrane protein that forms very stable ␤-sheet and can be processed into tape and other novel materials structures (33,34).…”

“…Several polymer-based peptide and protein self-assembly systems have also been developed by other investigators. These protein-based polymers were also designed de novo and produced through recombinant DNA biotechnology (35)(36)(37)(38)(39). Discovery and understanding of the nature of peptideand protein-based biological materials is an emerging field.…”

Applications of novel affinity cassette methods: use of peptide fusion handles for the purification of recombinant proteins

“…X a (McPherson et al, 1992) or exopeptidases such as dipeptidyl aminopeptidase-1 (DAP-1; Pedersen et al, 1999) or carboxypeptidase B . Whilst these enzymatic methods will generally act on the introduced accessible cleavage sequences, they have often been found to be less efficient since their kinetic rates of cleavage are affected by the accessibility of the cleavage site and the nature of the adjacent amino acid residues.…”