Conjugation of Amphiphilic Proteins to Hydrophobic Ligands in Organic Solvent
| Autor: | Harsha D. Magurudeniya, M. Lisa Phipps, Ryszard Michalczyk, Jennifer S. Martinez, Antonietta M. Lillo, Srinivas Iyer, Hung-Ju Yen, Reginaldo C. Rocha, Trideep Rajale, Eva Rose M. Balog, Timothy C. Sanchez, Ciana L Lopez, Hsing-Lin Wang |
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| Rok vydání: | 2018 |
| Předmět: |
Polymers
Pyridines Proton Magnetic Resonance Spectroscopy Biomedical Engineering Pharmaceutical Science Bioengineering 02 engineering and technology Conjugated system Ligands 010402 general chemistry 01 natural sciences Amphiphile Amines Organic Chemicals Solubility Pharmacology Ligand Chemistry Organic Chemistry Aqueous two-phase system 021001 nanoscience & nanotechnology Combinatorial chemistry Elastin 0104 chemical sciences Metals Covalent bond Solvents Electrophoresis Polyacrylamide Gel Spectrophotometry Ultraviolet Amine gas treating Protein Multimerization 0210 nano-technology Hydrophobic and Hydrophilic Interactions Biotechnology Conjugate |
| Zdroj: | Bioconjugate Chemistry. 29:2654-2664 |
| ISSN: | 1520-4812 1043-1802 |
| DOI: | 10.1021/acs.bioconjchem.8b00354 |
| Popis: | Protein-ligand conjugations are usually carried out in aqueous media in order to mimic the environment within which the conjugates will be used. In this work, we focus on the conjugation of amphiphilic variants of elastin-like polypeptide (ELP), short elastin (sEL), to poorly water-soluble compounds like OPPVs ( p-phenylenevinylene oligomers), triarylamines, and polypyridine-metal complexes. These conjugations are problematic when carried out in aqueous phase because hydrophobic ligands tend to avoid exposure to water, which in turn causes the ligand to self-aggregate and/or interact noncovalently with hydrophobic regions of the amphiphile. Ultimately, this behavior leads to low conjugation efficiency and contamination with strong noncovalent "conjugates". After exploring the solubility of sEL in various organic solvents, we have established an efficient conjugation methodology for obtaining covalent conjugates virtually free of contaminating noncovalent complexes. When conjugating carboxylated ligands to the amphiphile amines, we demonstrate that even when only one amine (the N-terminus) is present, its derivatization is 98% efficient. When conjugating amine moieties to the amphiphile carboxyls (a problematic configuration), protein multimerization is avoided, 98-100% of the protein is conjugated, and the unreacted ligand is recovered in pure form. Our syntheses occur in "one pot", and our purification procedure is a simple workup utilizing a combination of water and organic solvent extractions. This conjugation methodology might provide a solution to problems arising from solubility mismatch of protein and ligand, and it is likely to be widely applied for modification of recombinant amphiphiles used for drug delivery (PEG-antibodies, polymer-enzymes, food proteins), cell adhesion (collagen, hydrophobins), synthesis of nanostructures (peptides), and engineering of biocompatible optoelectronics (biological polymers), to cite a few. |
| Databáze: | OpenAIRE |
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