1. Electron transfer in biological systems

2. Biological electron transfer

3. Kinetics of electron transfer reactions
Electron transfer between 2 metal centers in metalloproteins is always via outer-sphere mechanism (no bridging ligand, coordination spheres remain essentially the same for both metal ions)
Reasonably fast (> 10 s-1) over large distances (up to 30 Å)
Can be rationalised by Marcus Theory (see Shriver/Atkins, 4th edition p. 516ff)

4. Simplified Marcus Theory
The reorganization energy l is defined as the energy required to “reorganize” the structure from initial to final coordinates, without changing the electronic state.
Free Energy
DG0#
DG0
Reaction coordinate: contains positions of atoms (nuclear coordinates)

5. Marcus Theory: Key points
For DG0 = - l , activation energy DG# becomes = 0: “activationless” e-transfer
Fast reactions if DG0 and l are similar to one another
there are “ideal” combinations of reaction enthalpy and reorganization energy
Often observed in biological systems: Small values for both

6. Cytochromes Fe-S proteins Blue copper proteins
e- transfer proteins
Cytochromes
Fe-S proteins
Blue copper proteins

7. Examples for efficient electron transfer units (1): Cytochromes
Name comes from the fact that they are coloured
Differ by axial ligands and whether covalently bound
Involved in electron transfer (a,b,c) or oxygen activation (P450)
Essential for many redox reactions

8. UV-Vis Spectra of cytochromes
classified by a bands:
a: nm
b: nm
c: nm
(there’s also d and f)
all involved in electron transfer, all CN6
P450: 450 nm:
Oxygen activation; CN5
Absorption spectra of oxidized (Fe(III)) and reduced (Fe(II)) horse cytochrome c.

9. Cytochrome c Small soluble proteins (ca. 12 kDa)
Near inner membrane of mitochondria
Transfers electrons between 2 membrane proteins ( for respiration)
Heme is covalently linked to protein via vinyl groups (thioether bonds with Cys)
1 Met and 1 His ligand (axial)
horse heart cytochrome c
Bushnell, G.W.,  Louie, G.V.,  Brayer, G.D. J.Mol.Biol. v pp , 1990
Conserved from bacteria to Man

10. Cytochromes b Heme has no covalent link to protein
Two axial His ligands
Shown is only soluble domain; the intact protein is bound to membrane
F Arnesano, L Banci, I Bertini, IC Felli: The solution structure of oxidized rat microsomal cytochrome b5. Biochemistry (1998) 37,

11. Why e- transfer in cytochromes is efficient
Porphyrin ring provides rigid scaffold: No significant changes in structure between Fe(II) and Fe(III) forms: relatively small reorganisation energy
Electron is delocalised over porphyrin ring: can be transferred efficiently over edge of ring

12. Not for electron transfer: the cytochromes P450 are oxygenases
CN5, axial ligand is a Cys
6th site for substrate/oxygen binding
Hydroxylates camphor
P450Cam

13. Tuning of heme function
In (deoxy)hemoglobin, Fe(II) is 5-coordinate
Must avoid oxidation to Fe(III) (Met-hemoglobin)
Neutral His ligand: His-Fe(II)-porphyrin is uncharged: Favourable
P450: Catalyses hydroxylation of hydrophobic substrates. Also 5-coordinate
1 axial Cys thiolate ligand (negatively charged): Resting state is Fe(III), also uncharged
In cytochromes, CN=6: No binding of additional ligand, but very effective 1 e- transfer
Neutral ligands (Met or His): Fe(II) more stabilised than Fe(III)

14. Examples for efficient electron transfer units (2): Fe-S proteins
Probably amongst the first enzymes
Generally, Fe (II) and (III), Cys thiolate and sulfide
Main function: fast e- transfer
At least 13 Fe-S clusters in mitochondrial respiration chain
Rubredoxins: mononuclear FeCys4 site
Ferredoxins: 2,3 or 4 irons

15. Rubredoxins: FeCys4
X-ray Structure of RUBREDOXIN from Desulfovibrio gigas at 1.4 A resolution.
FREY, M., SIEKER, L.C., PAYAN, F.

16. 1rfs: Spinach
Fe2S2(Cys-S)4
1 awd: CHLORELLA FUSCA
Fe2S2(Cys-S)2-(His-N)2: Rieske proteins
Fe3S4(Cys-S)4
Fe4S4(Cys-S)4
1fda: Azotobacter vinelandii

17. Fe-S clusters can be easily synthesised from Fe(III), sulfide and organic thiols, but are prone to rapid oxidation in air
Self-assembly of Fe-S clusters
Richard Holm

18. Delocalisation of electrons: Mixed valence
localized Fe3+ (red) and localized Fe2+ (blue) sites, and
delocalized Fe2.5+Fe2.5+ pairs (green)
Why e- transfer is fast:
Clusters can delocalize the “added” electron
minimizes bond length changes
decreases reorganization energy

19. Fe-S proteins often contain more than one cluster:
Fe-only hydrogenase from Clostridium pasteurianum
Activation of H2
Active site (binuclear Fe cluster) on top
The other five Fe-S clusters provide long-range electron transfer pathways
Pdb 1feh

20. Nitrogenase (Klebsiella pneumoniae)
Catalyses nitrogen fixation
P cluster
FeMoCo cofactor cluster
N2 + 8H+ + 8e ATP → 2NH3 + H2 + 16ADP + 16 Pi

21. Redox potentials

22. Tuning of redox potentials
For both heme proteins and Fe-S clusters, ligands coarsely tune redox potential
In [4Fe-4S] clusters, proteins can stabilise a particular redox couple through:
(a) solvent exposure of the cluster
(b) specific hydrogen bonding networks especially NH-S bonds
(c) the proximity and orientation of protein backbone and side chain dipoles
(d) the proximity of charged residues to the cluster

23. Tuning of redox potentials: Stabilisation of different redox states via weak interactions
Bacterial ferredoxins and HiPIPs: Both have Fe4S4Cys4 clusters
-400 mV vs mV
Ferredoxins: [Fe4S4Cys4]3- → [Fe4S4Cys4]2-
HiPIPs: [Fe4S4Cys4]2- → [Fe4S4Cys4]1-
HiPIPs are more hydrophobic: Favours -1
NH...S bonds: 8-9 in Fd, only 5 in HiPIPs
Compensate charge on cluster; -3 favoured
*) HiPIP: high potential iron-sulfur proteins

24. Examples for efficient e- -transfer (3): Blue copper proteins
Azurin, stellacyanin, plastocyanin
Unusual coordination geometry: Another example for how proteins tune metal properties
Consequences:
Curious absorption and EPR spectra
High redox potential (Cu(I) favoured)
No geometric rearrangement for redox reaction: Very fast

25. Blue copper proteins: coordination geometry
2.11 Å
2.9 Å
Angles also deviate strongly from ideal tetrahedron
(84-136°)
Amicyanin (pdb 1aac) from Paracoccus denitrificans

26. Key points Properties such as redox potentials are tuned by proteins
Coarse tuning by metal ligands
Charge imposed by ligand can favour particular oxidation state
Geometry can be imposed by protein: Can favour particular oxidation state, and also increase reaction rate
Fine tuning by “second shell”: hydrophobicity, hydrogen bonds, charges and dipoles in vicinity etc.