Solid phase peptide synthesis on epoxy-bearing methacrylate monoliths.
Monoliths based on a copolymer of glycidyl methacrylate (GMA) and ethylene dimethacrylate (EDMA) can be used directly as sorbents for affinity chromatography after solid phase peptide synthesis. The quality of the synthesized products, the amount of grown peptides on a support and the reproducibility of the process must be considered. A determination of the quantity of the introducing beta-Ala (and, consequently, the total amount of synthesized peptide) was carried out. Three peptides complementary to recombinant tissue plasminogen activator (t-PA) have been synthesized using Fmoc-chemistry on GMA-EDMA disks. The peptidyl ligands were analysed by amino acid analysis, ES-MS and HPLC methods. The affinity binding parameters were obtained from frontal elution data. The results were compared with those established for GMA-EDMA affinity sorbents formed by the immobilization of the same but separately synthesized and purified ligands. The immobilization on GMA-EDMA disks was realized using a one-step reaction between the amino groups of the synthetic ligand and the original epoxy groups of monolithic material. The affinity constants found for two kinds of sorbent did not vary significantly. Finally, the directly obtained affinity sorbents were tested for t-PA separation from a cellular supernatant.
Vlakh
E
E
Institute of Macromolecular Compounds, Russian Academy of Sciences.
Novikov
A
A
Vlasov
G
G
Tennikova
T
T
D002846:Chromatography, Affinity; D004852:Epoxy Compounds / Q000737:chemistry*; D005069:Evaluation Studies as Topic; D013058:Mass Spectrometry; D008689:Methacrylates / Q000737:chemistry*; D008722:Methods; D010455:Peptides / Q000138:chemical synthesis*; D010959:Tissue Plasminogen Activator / Q000737:chemistry / Q000302:isolation & purification; D015091:beta-Alanine / Q000737:chemistry
D004852:Epoxy Compounds; D008689:Methacrylates; D010455:Peptides; D015091:beta-Alanine; C004919:ethylene dimethacrylate; D010959:Tissue Plasminogen Activator; C007870:glycidyl methacrylate