title: Design and engineering of a man-made diffusive electron-transport protein.
abstract: Maquettes are man-made cofactor-binding oxidoreductases designed from first principles with minimal reference to natural protein sequences. Here we focus on water-soluble maquettes designed and engineered to perform diffusive electron transport of the kind typically carried out by cytochromes, ferredoxins and flavodoxins and other small proteins in photosynthetic and respiratory energy conversion and oxido-reductive metabolism. Our designs were tested by analysis of electron transfer between heme maquettes and the well-known natural electron transporter, cytochrome c. Electron-transfer kinetics were measured from seconds to milliseconds by stopped-flow, while sub-millisecond resolution was achieved through laser photolysis of the carbon monoxide maquette heme complex. These measurements demonstrate electron transfer from the maquette to cytochrome c, reproducing the timescales and charge complementarity modulation observed in natural systems. The ionic strength dependence of inter-protein electron transfer from 9.7×10(6) M(-1) s(-1) to 1.2×10(9) M(-1) s(-1) follows a simple Debye-Hückel model for attraction between +8 net charged oxidized cytochrome c and -19 net charged heme maquette, with no indication of significant protein dipole moment steering. Successfully recreating essential components of energy conversion and downstream metabolism in man-made proteins holds promise for in vivo clinical intervention and for the production of fuel or other industrial products. This article is part of a Special Issue entitled Biodesign for Bioenergetics--the design and engineering of electronic transfer cofactors, proteins and protein networks, edited by Ronald L. Koder and J.L. Ross Anderson.
authors.affiliation: Department of Biochemistry & Biophysics, Univ. of Pennsylvania, Philadelphia PA, USA.
authors.forename: Bryan A
authors.initials: BA
authors.lastname: Fry
authors.affiliation: Department of Biochemistry & Biophysics, Univ. of Pennsylvania, Philadelphia PA, USA.
authors.forename: Lee A
authors.initials: LA
authors.lastname: Solomon
authors.affiliation: Department of Biochemistry & Biophysics, Univ. of Pennsylvania, Philadelphia PA, USA.
authors.forename: P
authors.initials: P
authors.lastname: Leslie Dutton
authors.affiliation: Department of Biochemistry & Biophysics, Univ. of Pennsylvania, Philadelphia PA, USA. Electronic address: moserc@mail.med.upenn.edu.
authors.forename: Christopher C
authors.initials: CC
authors.lastname: Moser
mesh_terms: D000595:Amino Acid Sequence; D045304:Cytochromes c / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004058:Diffusion; D004579:Electron Transport / Q000235:genetics; D045222:Electron Transport Chain Complex Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006418:Heme / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010782:Photolysis; D015202:Protein Engineering / Q000379:methods*; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid
chemical_list: D045222:Electron Transport Chain Complex Proteins; D006418:Heme; D045304:Cytochromes c
keywords: Charge complementarity; Cytochrome c; Diffusion; Electron-transfer kinetics; Electrostatics; Inter-protein electron transfer; Man-made redox proteins; Maquettes; Protein design