Control of stability of polypeptide multilayer nanofilms by quantitative control of disulfidebond formation

Abstract

The crosslinking of polymers in a polymeric material will alter the mechanical properties ofthe material. Control over the mechanical properties of polyelectrolyte multilayer films(PEMs) could be useful for applications of the technology in medicine and other areas.Disulfide bonds are ‘natural’ polypeptide crosslinks found widely in wild-type proteins.Here, we have designed and synthesized three pairs of oppositely charged 32merpolypeptide to have 0, 4, or 8 cysteine (Cys) residues per molecule, and we havecharacterized physical properties of the peptides in a PEM context. The average lineardensity of free thiol in the designed peptides was 0, 0.125, or 0.25 per amino acid residue.The peptides were used to make 10-bilayer PEMs by electrostatic layer-by-layerself-assembly (LBL). Cys was included in the peptides to study specific effects of disulfidebond formation on PEM properties. Features of film assembly have been found todepend on the amino acid sequence, as in protein folding. Following polypeptideself-assembly into multilayer films, Cys residues were disulfide-crosslinked undermild oxidizing conditions. The stability of the crosslinked films at acidic pH hasbeen found to depend on the number of Cys residues per peptide for a givencrosslinking procedure. Crosslinked and non-crosslinked films have been analysed byultraviolet spectroscopy (UVS), ellipsometry, and atomic force microscopy (AFM) tocharacterize film assembly, surface morphology, and disassembly. A selective etchingmodel of the disassembly process at acidic pH is proposed on the basis of theexperimental data. In this model, regions of film in which the disulfide bonddensity is low are etched at a higher rate than regions where the density is high.