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Developing an Artificial Photosynthetic Device Christopher Hobbsa, Klaudia Wagnera, Pawel Wagnera, Keith Gordonc, Nicholas Roacha, Rhys a b b c b b Mitchell , Goutham Kodali , Bohdana Discher , Stasi Elliot , Christopher Mozer , P. Leslie Dutton a and David L. Officer aARC

Centre of Excellence for Electromaterials Science and the Intelligent Polymer Research Institute, University of Wollongong, Wollongong, NSW 2522, Australia bUniversity of Pennsylvania, Perelman School of Medicine, Stellar-Chance Labs, Suite 1005, 422 Curie Boulevard, Philadelphia, PA 19104-6059, USA cMacDiarmid Institute for Advanced Materials and Nanotechnology, Department of Chemistry, University of Otago, Dunedin 9054, New Zealand

Email: ch147@uowmail.edu.au

Introduction

Results and Discussion 0.50

Porphyrin N1 0.45 0.40

0.30 0.05

0.20 0.15 0.10

0.00

500

0.05

520

540

560

580

600

620

640

However, a much cleaner complexation occurs when a hydrophilic porphyrin (N2) is titrated against the BT6 maquette, a distinct ‘shift’ in the Soret peak of the porphyrin is observed (Fig 4). The two peaks at 433 and 423 nm correspond to the bound and unbound porphyrin monomers, respectively.

0.00 400

600

800

Porphyrin N2

1.0

0.8

0.6 433nm

0.045 0.040 0.035

0.4

0.030 0.025 0.020 0.015 0.010 0.005

0.2

0.000 -0.005 500

600

0.6

0.0 400

600

0.30

Purified Porphyirn/Maquette Porphyrin

423nm

0.25

0.2

Ltot*efree+(ebound-efree)*((Kd+Ptot+Ltot) -((Kd+Ptot+Ltot)^2-4*Ptot*Ltot)^0.5)/2

6.74911E-6

Reduced Chi-Sqr

0.99964

Adj. R-Squar

Value

0.0

D

absorbance (a.u)

0.4

Equation

0.20 433nm

0.15

0.10

Standard Err

efree

37177.8844 1588.00029

ebound

536676.950 6328.31073

[Maquette]

7.77814E-7

7.0587E-9

Calculated Kd

1.99834E-8

2.96023E-9

0.05

To isolate the pure maquetteporphyrin ensembles, a PD-10 size exclusion column is employed. Maquette BT6 is exposed to excess porphyrin N2 before being passed through the column. The elution is of pure maquette-porphyrin ensemble, as verified through UV-Vis. spectroscopy (Fig 6).

0.00

0.000000

0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2 2.1 2.2 2.3 2.4 2.5 2.6 2.7 2.8 2.9 3

800

Fig 4: A titration of a 0.5 µM BT6 maquette titrated with increments of 0.1 µM porphyrin N2.

Fig 3: A titration of a 0.5 µM BT6 maquette titrated with increments of 0.1 µM porphyrin N1.

The maquette-porphyrin titration (Fig 4) can be further analysed by evaluating the ‘bound’ porphyrin peak at 433 nm (Fig 5). The analysis shows a distinct inflection point which separates bound and unbound porphyrin. The inflection point allows for calculation of the dissociation constant, Kd, a direct measure of binding affinity, here determined to be 20 nM.

423nm

wavelenght (nm)

Wavelength (nm)

absorbance (a.u)

absorbance (a.u)

0.35

0.25

Maquettes readily bind 1 or 2 porphyrin cofactors, depending on the availability of histidines. When a hydrophobic porphyrin (N1) is titrated against BT6 maquette in aqueous buffer, an increase in a ‘bound’ state of the porphyrin Soret peak is observed (Fig 3), with the formation of porphyrin aggregates in solution.

0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1 2.2 2.3 2.4 2.5 2.6 2.7 2.8 2.9 3.0

432nm

Fig 2: Schematic representation of an artificial photosynthetic system for water splitting.

absorbance (a.u)

Fig 1: A representation of a ‘maquette’ with two porphyrins.

One of the problems facing the 21st Century is the alarming rate at which fossil fuel reserves are diminishing, coupled with the rate of increase in energy demands. New ways to produce clean renewable sources of energy are urgently needed. An ‘artificial photosynthetic device’ could potentially reproduce some of the processes in natural photosynthesis, converting sunlight into energy (electricity) or a solar fuel such as hydrogen. The challenge in developing an artificial photosynthetic device is to use known, readily available materials, such that the process of photosynthesis is simplified. Dutton et al.1 has developed de novo synthetically designed protein scaffolds, known as maquettes (Fig 1), capable of incorporating cofactors such as hemes, flavins, and porphyrins via histidine ligation2.

Incorporation of porphyrins into maquettes would represent a path to the development of an artificial photosynthetic reaction centre (Fig 1). This would incorporate a porphyrin antenna system for light capture, transferring the excited energy to a maquette reaction centre (RC), coupled to a water oxidising catalyst (WOC) and a proton reducing catalyst (PRC) on a suitable electrode. Excitation of the antenna would lead to electricity generation and H2 production (Fig 2).

0.000001

0.000002

0.000003

Porphyrin Concentration (M)

Fig 5: Analysis of the BT6/N2 binding titration shown in Fig 4.

Future Work The binding porphyrins into maquettes is a fundamental step towards developing an artificial photosynthetic device. Future endeavours towards this ultimate goal include:  Binding dimeric and trimeric porphyrins  Covalently attaching the maquette-porphyrin ensemble onto a substrate/electrode  Coupling catalysts to the electrode-bound maquette-porphyrin ensemble.

This work represents the first wavelength (nm) steps towards the development an efficient artificial Fig 6: UV-vis. spectra of pure porphyrin and BT6/N2 of ensemble. photosynthetic device. 400

600

800

References 1- Dutton, P. L.; Moser, C. C. Engineering Enzymes. Farday Discussion (2011) 148, 443-448. 2- Gibney, B. R.; Isogai, Y.; Rabanal, F.; Reddy, K. S.; Grosset, A. M.; Moser, C. C.; Dutton, P. L. Self Assembly of Heme A and Heme B in a Designed Four-Helix Bundle: Implications for a Cytochrome c Oxidase Maquette. Biochemistry (2000) 39, 11041-11049.


Chris hobbs