title: On-surface assembly of coiled-coil heterodimers.
abstract: The coiled coil is a widespread protein motif responsible for directing the assembly of a wide range of protein complexes. To date, research has focused largely on the solution phase assembly of coiled-coil complexes. Here, we describe an investigation into coiled-coil heterodimer assembly where one of the peptides is immobilized directly onto a gold electrode. Immobilization is achieved by the introduction of a unique cysteine residue at the C terminus, allowing for covalent and orientated attachment to a thiol-reactive surface, here the gold electrode. We show an electrochemical impedance of the resulting self-assembled polypeptide monolayer around |Z| = 4 × 10(4) Ω cm(2) at 100 mHz with a minimum phase angle of -84°, consistent with the formation of a densely packed, insulating layer. The thickness of the peptide monolayer, as measured using ellipsometry, is around 3 nm, close to that expected for a self-assembled monolayer assembled from helical peptides. Crucially, we find that the efficiency of dimerization between a peptide in solution and its coiled-coil partner peptide immobilized on a surface is strongly dependent upon the density of the immobilized peptide layer, with dimer assembly being strongly suppressed by high-density peptide monolayers. We thus develop an approach for controlling the density of the immobilized peptide by diluting the monolayer with a thiolated, random-coil peptide to modulate dimerization efficiency and demonstrate electrochemical detection of highly specific, coiled-coil heterodimer on-surface assembly.
authors.affiliation: Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds, United Kingdom.
authors.forename: Simon J
authors.initials: SJ
authors.lastname: White
authors.forename: D William A
authors.initials: DW
authors.lastname: Morton
authors.forename: Boon Chong
authors.initials: BC
authors.lastname: Cheah
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Bronowska
authors.forename: A Giles
authors.initials: AG
authors.lastname: Davies
authors.forename: Peter G
authors.initials: PG
authors.lastname: Stockley
authors.forename: Christoph
authors.initials: C
authors.lastname: Wälti
authors.forename: Steven
authors.initials: S
authors.lastname: Johnson
mesh_terms: D019281:Dimerization; D055664:Electrochemical Techniques; D004566:Electrodes; D006046:Gold / Q000737:chemistry; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry*; D013438:Sulfhydryl Compounds / Q000737:chemistry; D013499:Surface Properties
chemical_list: D010455:Peptides; D013438:Sulfhydryl Compounds; D006046:Gold