1 research outputs found
Conjugation of Amphiphilic Proteins to Hydrophobic Ligands in Organic Solvent
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 (<i>p</i>-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