The effect of a macrocyclic constraint on electron transfer in helical peptides: A step towards tunable molecular wires

Article abstract of DOI:10.1039/c3cc47885h

Two helical peptides, one constrained by a covalent side-chain staple, exhibit vastly different electronic properties despite adopting essentially the same backbone conformation. High level calculations confirm that these differences are due to the additional backbone rigidity imparted by the macrocyclic constraint.

Full text of DOI:10.1039/c3cc47885h

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The effect of a macrocyclic constraint on electron
transfer in helical peptides: A step towards tunable
molecular wires†
Cite this: Chem. Commun., 2014,
50, 1652
Received 14th October 2013,
Accepted 9th December 2013
Jingxian Yu,*a John R. Horsley,a Katherine E. Moore,b Joe G . Shapterb and
Andrew D. Abell*a
DOI: 10.1039/c3cc47885h
www.rsc.org/chemcomm
Two helical peptides, one constrained by a covalent side-chain The peptides were designed to share a common 310-helical back-
staple, exhibit vastly different electronic properties despite adopting bone conformation, with peptide 1 possessing a rigid backbone
essentially the same backbone conformation. High level calculations conformation as defined by the side-chain constraint (Scheme 1).
confirm that these differences are due to the additional backbone
rigidity imparted by the macrocyclic constraint.
The geometries of peptides 1 and 2 were confirmed as 310-helical
by H NMR spectroscopy. In particular, NH (i) to NH (i + 1) ROESY
1
correlations were observed for both peptides and also their synthetic
Helical domains in peptides and proteins provide a good medium precursors, 3–5 and 6 and 7. Furthermore, CaH (i) to NH (i + 1) and
for electron transfer1–5 over surprisingly long molecular distances medium range CaH (i) to NH (i + 2) were found for 1 and 2, with long
(>100 Å).6 Such structures present as ideal candidates for use as range CaH (i) to NH (i + 3) correlations evident in peptides 3, 5 and 7,
molecular wires, particularly when combined with an ability to be as detailed in the ESI.† A lack of CaH (i) to NH (i + 4) correlations for
precisely functionalised.7–9 However, more detailed understanding all peptides precludes the possibility of an a-helical structure, which is
of exactly what defines and controls the mechanisms and characterised by (i to i + 4) hydrogen bonds.18 A strong negative
efficiency of electron transfer in peptides is required before minimum near 202 nm, with a far weaker minimum at approximately
this promise can be fully realised. Toward this goal, we recently 232 nm was observed in a representative CD spectrum of the con-
demonstrated that intramolecular hydrogen bonding within helical strained peptide 3 (see ESI†), which further supports a 310-helical
Aib (a-aminoisobutyric acid) containing oligomers plays a critical conformation.19,20 Collectively, this information confirms the presence
role in defining the mechanism of electron transfer.10,11 Recent of a 310-helical backbone structure and that the C-terminal ferrocene
theoretical studies suggest that low-frequency rotation between and the constraint do not impinge on the backbone helicity.
neighbouring amino acids brings adjacent carbonyl groups into
The lowest energy conformers for the N-protected peptides 5
alignment to allow efficient charge transfer through the peptide.12,13 and 7 were determined by molecular modelling (using a hybrid
As such any feature that enhances backbone rigidity should restrict B3LYP method with the 6-31G** basis set for all C, H, N, O atoms,
molecular motion and consequently retard electron transfer in
peptides. However, the exact influence of backbone rigidity on
electron transfer requires significant further investigation.14–17
Here we present electrochemical and theoretical studies on a
helical peptide constrained by a side-chain tether to begin to
unravel these effects in isolation from other factors such as
chain length, dipole orientation and the associated hydrogen
bonding that are known to influence electron transfer.
The required Aib-rich hexapeptide stapled with an i to i + 3
macrocyclic constraint by Huisgen cycloaddition (peptide 1) and a
linear analog (peptide 2) were synthesised as described in ESI.†
a School of Chemistry and Physics, The University of Adelaide, Adelaide, SA 5005,
Australia. E-mail: jingxian.yu@adelaide.edu.au, andrew.abell@adelaide.edu.au
b School of Chemical & Physical Sciences, Flinders University, Bedford Park,
SA 5042, Australia
Scheme 1 Structures of target peptides 1 and 2 and key synthetic
† Electronic supplementary information (ESI) available. See DOI: 10.1039/c3cc47885h
intermediates.
1652 | Chem. Commun., 2014, 50, 1652--1654
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and the Lanl2dz basis set for Fe atoms) in order to further define
Despite having very similar backbone geometries, the two
the backbone conformations of the constrained and unconstrained target peptides exhibit considerably different formal potentials
peptides. The N-protected peptides were used in these studies as free and electron transfer rate constants. Side-bridge stapling as in
amines are known to give rise to unrealistic electrostatic interactions, peptide 1 results in a significant formal potential shift to the
resulting in unstable lowest energy conformers.21 The resulting positive of approximately 480 mV in comparison to peptide 2.
structures (see ESI†) reveal that the distances from the first to last Thus oxidation–reduction of the redox-active ferrocene moiety in
carbonyl carbons (backbone lengths) of the two peptides are similar, the constrained peptide is energetically much less favourable than
11.94 Å and 12.02 Å for peptides 5 and 7 respectively. In addition, the in the linear analog. Such a dramatic formal potential shift has
greatest variation between hydrogen bond lengths is 0.14 Å. The not, to the best of our knowledge, been previously reported in
mean dihedral angles for residues 1–4 of peptide 5 were calculated to ferrocene-derivatised peptides. As both peptides essentially differ
be À56.861 for Ø and À31.291 for C, deviating from an ideal 310-helix only in the presence (or absence) of the side-bridge constraint, the
by 0.141 and 1.291, respectively,22 whilst the mean dihedral angles for significant formal potential shift found in 1 is clearly a result of
residues 1–4 deviated from an ideal 310-helix by 1.331 and 3.751 in additional backbone rigidity imparted by this constraint. Our
peptide 7. These studies demonstrate that the two structures have experimental data also reveal a significant decrease in the electron
essentially the same conformations, such that they differ only in the transfer rate constant (approx. 25%) upon introducing the con-
presence (or absence) of the constraint and the associated effect that straint of 1. Previous studies have shown that electron transfer
this has on backbone rigidity.
rate constants in peptides can vary greatly,6,10,17 but without such
The constrained peptide 1 and linear analog 2 were next a significant difference in formal potential as reported here. Our
separately attached to vertically aligned single-walled carbon results suggest that side-bridge stapling creates an additional
nanotube array/gold (SWCNTs/Au) electrodes23,24 (see ESI† for reorganisation energy barrier that impedes electron transfer
details) in order to study their electron transfer kinetics. SWCNTs/ within the peptide, in turn decreasing the charge transfer rate.
Au electrodes provide a high surface concentration of attached Hence reducing the backbone flexibility within a helical peptide
redox probes and hence high sensitivity and reproducibility of through the introduction of a constraint lowers the rate of
electrochemical measurement.25–27 Fig. 1 shows the cyclic electron transfer by restricting the precise torsional motions that
voltammograms obtained for the two target peptides immersed in lead to facile intramolecular electron transfer along the backbone.
0.1 mol LÀ1 TBAPF6–CH3CN solutions. These show a pair of redox Thus side-bridge stapling provides a unique approach to mani-
peaks, characteristic of a one-electron oxidation–reduction reaction pulate energy barriers and conductance in peptides.
(Fc+/Fc). The surface concentrations of the peptides were deter-
Interestingly, the all Aib containing linear peptide 8 (see ESI†)
mined by integrating background current subtracted peak areas to gave a formal potential shift to the positive of 137 mV and approxi-
be 3.76 Â 10À10 mol cmÀ2 for 1 and 2.52 Â 10À10 mol cmÀ2 for 2 mate two-fold decrease in the electron transfer rate constant
(see Table 1). These surface concentrations are comparable to other compared to 2. This peptide adopts essentially the same backbone
carbon nanotube electrode studies.10,23 The formal potentials conformation as the earlier peptides based on NMR and modelling
(Eo) and apparent electron transfer rate constants (kapp) were studies as discussed in the ESI.† However, it would be expected to
estimated to be 0.853 V and 28.1 sÀ1 for 1 and 0.371 V and have somewhat reduced backbone flexibility relative to 2 with the
117.3 sÀ1 for 2, respectively (as detailed in Table 1), using inclusion of an additional geminally disubstituted residue. The
Laviron’s formalism.28
electrochemical data are consistent with this notion, where
the electron transfer rate constant is intermediary between that of
the constrained peptide 1 and its linear analog 2. This observation
further supports the link between backbone flexibility and the rate
of electron transfer, as has also been noted for DNA and PNA.29,30
Theoretical calculations, using the latest constrained density
functional theory (cDFT),31 were performed on the model peptides
9 and 10 (see Fig. 2) in order to corroborate our experimental
observations. These peptides were chosen for this study since they
contain the same sequence as 1 and 2, but with ferrocene units at
both termini to act as both a donor and an acceptor. Calculation
of reorganisation energies for electron transfer along the back-
bone then provides an insight into the intramolecular electron
transfer dynamics. Diabatic states were constructed by indivi-
dually localising an overall charge of 1 on each of the ferrocene
units and amino acids, as shown in Fig. 2. Diabatic potential
profiles were determined by assuming that during an electron
hopping step, the nuclear configuration changes smoothly
between the optimised geometries of the diabatic states in which
the excess charge is localised before and after electron transfer32
Fig. 1 (a) Cyclic voltammograms of peptide 1 (blue) and peptide 2 (red)
immobilised on SWCNTs/Au electrodes taken at 5 V sÀ1. (b) Peak potential
versus ln(scan rate) for peptides 1 and 2 after background current subtraction.
Table 1 Electron transfer rate constants (kapp), surface concentrations
and formal potentials (Eo) of the constrained peptide (peptide 1) and linear
analog (peptide 2)
Surface concentration
Eo
kapp
(Â10À10 mol cmÀ2
)
(V vs. AgCl/Ag)
(sÀ1
)
Peptide
1
2
3.76 Æ 0.35
0.853
0.371
28.1 Æ 3.6
2.52 Æ 0.18
117.3 Æ 9.9 (see ESI†). Peptides 9 and 10 show comparable reorganisation
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electron transfer by enhancing the rigidity of the peptide back-
bone. This then generates an additional reorganisation energy
barrier which restricts the necessary torsional motions that lead to
facile intramolecular electron transfer along the backbone. Our
results provide definitive evidence of a direct link between back-
bone rigidity and electron transfer in peptides. These findings
provide a new means to fine tune the rates of electron transfer in
peptides, which represent an important step towards their imple-
mentation into molecular electronic assemblies.
Notes and references
1 H. S. Mandal and H.-B. Kraatz, J. Phys. Chem. Lett., 2012, 3, 709.
2 R. A. Malak, Z. N. Gao, J. F. Wishart and S. S. Isied, J. Am. Chem. Soc.,
2004, 126, 13888.
3 P. A. Brooksby, K. H. Anderson, A. J. Downard and A. D. Abell,
J. Phys. Chem. C, 2011, 115, 7516.
4 J. Yu, D. M. Huang, J. G. Shapter and A. D. Abell, J. Phys. Chem. C,
2012, 116, 26608.
5 Y.-F. Wang, Z.-Y. Yu, J. Wu and C.-B. Liu, J. Phys. Chem. A, 2009, 113,
10521–10526.
Fig. 2 Constructed diabatic states in constrained peptide 9 (top) and
unconstrained peptide 10 (bottom). Charge localisation fragments of the
molecule involving the side bridge are indicated using two different
colours in peptide 9.
6 Y. Arikuma, H. Nakayama, T. Morita and S. Kimura, Angew. Chem.,
Int. Ed., 2010, 49, 1800.
7 J. A. Gao, P. Muller, M. Wang, S. Eckhardt, M. Lauz, K. M. Fromm
and B. Giese, Angew. Chem., Int. Ed., 2011, 50, 1926.
8 J. Watanabe, T. Morita and S. Kimura, J. Phys. Chem. B, 2005,
109, 14416.
9 M. Lauz, S. Eckhardt, K. M. Fromm and B. Giese, Phys. Chem. Chem.
Phys., 2012, 14, 13785–13788.
Table 2 Comparison of computed reorganisation energies for electron
hopping steps involving diabatic state S3 in the two model peptides (9 and 10)
Hopping
step
Peptide 9
Peptide 10
Difference
(kcal molÀ1
)
(kcal molÀ1
)
(kcal molÀ1
)
S2 - S3
S3 - S2
S3 - S4
S4 - S3
28.99
30.62
25.41
30.68
24.09
24.57
22.27
23.71
4.90
6.05
3.14
6.97
10 J. Yu, O. Zvarec, D. M. Huang, M. A. Bissett, D. B. Scanlon, J. G.
Shapter and A. D. Abell, Chem. Commun., 2012, 48, 1132.
11 J. Yu, J. R. Horsley and A. D. Abell, Aust. J. Chem., 2013, 66, 848.
12 E. W. Schlag, S. Y. Sheu, D. Y. Yang, H. L. Selzle and S. H. Lin, Proc.
Natl. Acad. Sci. U. S. A., 2000, 97, 1068.
13 E. W. Schlag, S. Y. Sheu, D. Y. Yang, H. L. Selzle and S. H. Lin, Angew.
Chem., Int. Ed., 2007, 46, 3196.
14 S. Okamoto, T. Morita and S. Kimura, Langmuir, 2009, 25, 3297.
15 G. A. Orlowski, S. Chowdhury and H. B. Kraatz, Electrochim. Acta,
2007, 53, 2034.
16 K. Takeda, T. Morita and S. Kimura, J. Phys. Chem. B, 2008, 112, 12840.
17 M. M. Galka and H. B. Kraatz, ChemPhysChem, 2002, 3, 356.
18 Z. Biron, S. Khare, A. O. Samson, Y. Hayek, F. Naider and J. Anglister,
Biochemistry, 2002, 41, 12687.
19 C. Toniolo, A. Polese, F. Formaggio, M. Crisma and J. Kamphuis,
J. Am. Chem. Soc., 1996, 118, 2744.
20 A. K. Boal, I. Guryanov, A. Moretto, M. Crisma, E. L. Lanni, C. Toniolo,
R. H. Grubbs and D. J. O’Leary, J. Am. Chem. Soc., 2007, 129, 6986.
21 N. A. Burton, M. J. Harrison, J. C. Hart, I. H. Hillier and D. W. Sheppard,
Faraday Discuss. Chem. Soc., 1998, 110, 463.
22 O. Jacobsen, H. Maekawa, N. H. Ge, C. H. Gorbitz, P. Rongved,
O. P. Ottersen, M. Amiry-Moghaddam and J. Klaveness, J. Org. Chem.,
2011, 76, 1228.
23 J. J. Gooding, R. Wibowo, J. Q. Liu, W. R. Yang, D. Losic, S. Orbons,
F. J. Mearns, J. G. Shapter and D. B. Hibbert, J. Am. Chem. Soc., 2003,
125, 9006.
24 K. E. Moore, B. S. Flavel, A. V. Ellis and J. G. Shapter, Carbon, 2011, 49, 2639.
25 K. E. Moore, B. S. Flavel, J. Yu, A. D. Abell and J. G. Shapter,
Electrochim. Acta, 2013, 89, 206.
energies for all electron hopping steps, except those involving
diabatic state S3. The reorganisation energies for the forward and
backward electron hopping steps from diabatic state S3 in 9 are
much higher than those for the corresponding steps in 10
(see Table 2). The introduction of the side-bridge gives rise to a
significant increase in reorganisation energy, in the range of
3.14–6.97 kcal molÀ1. Thus the higher reorganisation energy
barrier in peptide 9 is a direct result of the side-bridge constraint,
thus further supporting our experimental results.
In summary, electrochemical studies are reported on two
peptides containing Aib residues that constrain the backbones
into a well-defined 310-helix, a secondary structure known to favour
electron transfer. The first of these peptides (1) has its helical
geometry stabilised with a covalent constraint that links its i and
i + 3 side chains, resulting in additional conformational rigidity in
the backbone. Electrochemical studies revealed that peptide 1
exhibited a significant formal potential shift to the positive
(480 mV), and a substantial decrease in the electron transfer rate 26 K. T. Constantopoulos, C. J. Shearer, A. V. Ellis, N. H. Voelcker and
J. C. Shapter, Adv. Mater, 2010, 22, 557.
27 P. Diao and Z. F. Liu, Adv. Mater., 2010, 22, 1430.
28 E. Laviron, J. Electroanal. Chem., 1979, 100, 263.
constant (25%), compared to the unconstrained peptide 2. These
differences reflect the extent of backbone rigidity imparted by the
side-bridge constraint. In support, the all Aib containing linear 29 E. Wierzbinski, A. de Leon, X. Yin, A. Balaeff, K. L. Davis, S. Reppireddy,
R. Venkatramani, S. Keinan, D. H. Ly, M. Madrid, D. N. Beratan,
C. Achim and D. H. Waldeck, J. Am. Chem. Soc., 2012, 134, 9335.
30 E. Hatcher, A. Balaeff, S. Keinan, R. Venkatramani and D. N. Beratan,
peptide 8 displayed formal potential and electron transfer rate
constant values between those of peptides 1 and 2. This reflects an
intermediary backbone rigidity for this peptide. High level calcula-
tions confirm that the additional reorganisation energy barrier is a
direct result of the backbone rigidity imparted by the side-bridge
constraint. Thus the tether significantly impedes intramolecular
J. Am. Chem. Soc., 2008, 130, 11752.
31 T. Van Voorhis, T. Kowalczyk, B. Kaduk, L. P. Wang, C. L. Cheng and
Q. Wu, Annu. Rev. Phys. Chem., 2010, 61, 149–170.
32 A. Farazdel, M. Dupuis, E. Clementi and A. Aviram, J. Am. Chem.
Soc., 1990, 112, 4206.
1654 | Chem. Commun., 2014, 50, 1652--1654
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