title: Nanoscopic and redox characterization of engineered horse cytochrome C chemisorbed on a bare gold electrode.
abstract: In this paper, we exploit the potential offered by site-directed mutagenesis to achieve direct adsorption of horse cyt c on a bare gold electrode surface. To this issue, the side chain T102 has been replaced by a cysteine. T102 is close to the surface exposed C-terminal residue (E104), therefore the T102C mutation is expected to generate an exposed cysteine side chain able to facilitate protein binding to the electrode via the sulphur atom (analogously to what observed for yeast iso-1-cyt c). Scanning Tunnelling and Tapping Mode Atomic Force Microscopy measurements show that the T102C mutant stably adsorbs on an Au(111) surface and retains the morphological characteristics of the native form. Cyclic voltammetry reveals that the adsorbed variant is electroactive; however, the heterogeneous electron transfer with the electrode surface is slower than that observed for yeast iso-1-cyt c. We ascribe it to differences in the tertiary architecture of the two proteins, characterized by different flexibility and stability. In particular, the region where the N- and C-terminal helices get in contact (and where the mutation occurs) is analyzed in detail, since the interactions between these two helices are considered crucial for the stability of the overall protein fold.
authors.affiliation: Biophysics and Nanoscience Centre, CNISM, Dipartimento di Scienze Ambientali, Università della Tuscia, 01100 Viterbo, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Andolfi
authors.forename: Paola
authors.initials: P
authors.lastname: Caroppi
authors.forename: Anna Rita
authors.initials: AR
authors.lastname: Bizzarri
authors.forename: Maria Cristina
authors.initials: MC
authors.lastname: Piro
authors.forename: Federica
authors.initials: F
authors.lastname: Sinibaldi
authors.forename: Tommaso
authors.initials: T
authors.lastname: Ferri
authors.forename: Fabio
authors.initials: F
authors.lastname: Polticelli
authors.forename: Salvatore
authors.initials: S
authors.lastname: Cannistraro
authors.forename: Roberto
authors.initials: R
authors.lastname: Santucci
mesh_terms: D000818:Animals; D003545:Cysteine / Q000737:chemistry; D045304:Cytochromes c / Q000737:chemistry* / Q000378:metabolism; D004563:Electrochemistry / Q000379:methods; D004566:Electrodes; D006046:Gold / Q000737:chemistry*; D006736:Horses; D018625:Microscopy, Atomic Force; D016252:Microscopy, Scanning Tunneling / Q000379:methods; D008968:Molecular Conformation; D036103:Nanotechnology / Q000379:methods; D010084:Oxidation-Reduction; D015202:Protein Engineering; D017510:Protein Folding; D017434:Protein Structure, Tertiary
chemical_list: D006046:Gold; D045304:Cytochromes c; D003545:Cysteine