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Increasing Efficiency of Photoelectronic Conversion by Encapsulation of Photosynthetic Reaction Center Proteins in Arrayed Carbon Nanotube Electrode Nikolai Lebedev,*,† Scott A. Trammell,† Stanislav Tsoi,† Anthony Spano,‡ Jin Ho Kim,§ Jimmy Xu,§ Mark E. Twigg,| and Joel M. Schnur⊥ Center for Bio/Molecular Science and Engineering, Code 6900, U.S. NaVal Research Laboratory, Washington, D.C. 20375, Department of Biology, UniVersity of Virginia, CharlottesVille, Virginia 22901, DiVision of Engineering, Brown UniVersity, ProVidence, Rhode Island 02912, Electronic Science and Technology DiVision, Code 6812, U.S. NaVal Research Laboratory, Washington, D.C. 20375, and College of Science, George Mason UniVersity, Fairfax, Virginia 22030 ReceiVed April 10, 2008. ReVised Manuscript ReceiVed May 29, 2008 The construction of efficient light energy converting (photovoltaic and photoelectronic) devices is a current and great challenge in science and technology and one that will have important economic consequences. Here we show that the efficiency of these devices can be improved by the utilization of a new type of nano-organized material having photosynthetic reaction center proteins encapsulated inside carbon nanotube arrayed electrodes. In this work, a generically engineered bacterial photosynthetic reaction center protein with specifically synthesized organic molecular linkers were encapsulated inside carbon nanotubes and bound to the inner tube walls in unidirectional orientation. The results show that the photosynthetic proteins encapsulated inside carbon nanotubes are photochemically active and exhibit considerable improvement in the rate of electron transfer and the photocurrent density compared to the material constructed from the same components in traditional lamella configuration.
Introduction The harvesting and subsequent conversion of light energy is generally much more efficient in natural systems than in manmade ones. However, natural photoconversion processes are meant to be internal. As such, they present a rather difficult challenge to the effort of harvesting energy via natural processes for generating electrical power. The challenge is rooted in how to efficiently interface the internal photosynthetic units of proteins with the external electrodes and do so without external intervention. The construction of an efficient electrical conduit that interfaces natural photosynthetic proteins with man-made electrodes requires major advances in both protein engineering and electrical and material engineering at the proteins level, i.e., the nanoscale. The most efficient photovoltaic materials gifted by nature are the photosynthetic reaction center (RC) proteins. They have quantum conversion efficiency equal to 1 and can generate power conversion efficiency up to 60%.1–3 The principles of charge separation in photosynthetic RC are successfully used for the construction of organic photoactive charge separating complexes.4–6 Moreover, the direct utilization of photosynthetic RC in artificial or hybrid devices can serve as a paradigm for the component assembly in photovoltaic devices that substantially * To whom correspondence should be addressed. E-mail: nnl@ cbmse.nrl.navy.mil. † Center for Bio/Molecular Science and Engineering, Code 6900, U.S. Naval Research Laboratory. ‡ University of Virginia. § Brown University. | Electronic Science and Technology Division, Code 6812, U.S. Naval Research Laboratory. ⊥ George Mason University. (1) Hoff, A. J.; Deisenhofer, J. Phys. Rep.-ReV. Sect. Phys. Lett. 1997, 287, 2–247. (2) Sundstrom, V. Progress Quant. Electron. 2000, 24, 187–238. (3) Engel, G. S.; Calhoun, T. R.; Read, E. L.; Ahn, T. K.; Mancal, T.; Cheng, Y. C.; Blankenship, R. E.; Fleming, G. R. Nature 2007, 446, 782–786.
improves their performance.7–10 A prerequisite for utilizing the RC protein in artificial bioinorganic devices is the immobilization of the protein on an electrode surface while retaining its natural function. Moreover, to maximize efficiency most or all protein molecules must have the same desired orientation to the electrode. This can be achieved by using genetically engineered bacterial photosynthetic reaction centers and specifically synthesized organic linkers. Indeed, using these approaches we were able to construct monomolecular layers of aligned bacterial photosynthetic RC proteins on surfaces gold, glass, ITO, and carbon electrodes.11,12 Our experiments have shown that after binding to an electrode, photosynthetic RC can undergo efficient photo induced charge separation, operate as a photo rectifier and transfer current in only one direction consistent with the orientation of the protein.11,12 Carbon nanotubes (CNT) are very interesting nanomaterials that are size-compatible with proteins and physically accessible to proteins on both the exterior and interior tube wall-surfaces. They are known to exhibit superior and unique electron transfer (ET), thermal and mechanical properties. The possibility of CNT utilization for electronic, sensing and signaling devices has already (4) Hasobe, T.; Fukuzumi, S.; Kamat, P. V. J. Phys. Chem. B 2006, 110, 25477–25484. (5) D’Souza, F.; Chitta, R.; Sandanayaka, A. S. D.; Subbaiyan, N. K.; D’Souza, L.; Araki, Y.; Ito, O. J. Am. Chem. Soc. 2007, 129, 15865–15871. (6) Chitta, R.; Sandanayaka, A. S. D.; Schumacher, A. L.; D’Souza, L.; Araki, Y.; Ito, O.; D’Souza, F. J. Phys. Chem. C 2007, 111, 6947–6955. (7) Page, C. C.; Moser, C. C.; Chen, X. X.; Dutton, P. L. Nature 1999, 402, 47–52. (8) Lebedev, N.; Trammell, S. A.; Spano, A.; Lukashev, E.; Griva, I.; Schnur, J. J. Am. Chem. Soc. 2006, 128, 12044–12045. (9) Trammell, S. A.; Griva, I.; Spano, A.; Tsoi, S.; L.M., T.; Schnur, J.; Lebedev, N. J. Phys. Chem. B 2008, in press. (10) Willner, I. Science 2002, 298, 2407–2408. (11) Trammell, S. A.; Spano, A.; Price, R.; Lebedev, N. Biosens. Bioelectron. 2006, 21, 1023–1028. (12) Trammell, S. A.; Wang, L. Y.; Zullo, J. M.; Shashidhar, R.; Lebedev, N. Biosens. Bioelectron. 2004, 19, 1649–1655.
10.1021/la8011348 CCC: $40.75 2008 American Chemical Society Published on Web 07/11/2008
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been demonstrated.13–22 One feature that is under-explored so far is that CNT electrodes have an extended surface area that promises to provide an additional advantage for chemical to electronic signal transduction. Among CNT electrodes, the most attractive forms for our purpose are aligned CNT arrays. Precise size and spatial organization of each tube in these arrays allow for specific individually addressable manipulation.17,20,23 The possibility of CNT functionalization by proteins, nanoparticles and even single molecules bound to their tube wall surface has been demonstrated.24–26 However, up to now the functionalization was performed mainly by binding these compounds to exterior wall surfaces of CNT. Meanwhile, the encapsulation of foreign compounds inside CNT can lead to the construction of new nanoorganized material with high functional density, increased stability, one-dimensional mass and electron transfer. Nevertheless, the encapsulation of organic molecules, metal nanoparticles, and other small carbon nanostructures, like fullerenes, inside CNT was demonstrated with more limited successes and in small quantity (often at the level of individual tubes sporadically distributed in a solution). These new types of so-called “X@CNT” materials, show unique properties, some of which completely different from those observed with same components attached to the outer CNT walls.27–33 The present work represents the first report on the attempt to encapsulate and retain the function of relatively large multisubunit protein structures, like photosynthetic proteins, inside CNT.
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Figure 1. Scanning electron micrograph of the top of CNT array used in these experiments. The average diameter of the tube is 50-60 nm; the average distance between the tube centers is 100-110 nm.
Experimental Section The succinimidyl ester of 1-pyrenebutanoic acid was purchased from Molecular Probes (Eugene, OR). 2,3-Dimethoxy-5-methyl6-geranyl-1,4-benzoquinone (ubiquinone-2, Q2) and horse heart cytochrome c were purchased from Sigma. Highly ordered pyrolytic graphic (HOPG) was obtained from SPI Supplies. HOPG and CNT electrodes were fabricated by attaching a silver wire to the backside of the carbon substrate using silver epoxy (Epoxy Technology, MA). The backing and silver wire were insulated from the electrolyte by the application of an insulating epoxy (Epoxy Technology, MA). In the case of HOPG electrodes, a (13) Allen, B. L.; Kichambare, P. D.; Star, A. AdV. Mater. 2007, 19, 1439– 1451. (14) Bandaru, P. R. J. Nanosci. Nanotechnol. 2007, 7, 1239–1267. (15) Mahar, B.; Laslau, C.; Yip, R.; Sun, Y. IEEE Sensors J. 2007, 7, 266–284. (16) Avouris, P.; Chen, J.; Freitag, M.; Perebeinos, V.; Tsang, J. C. Phys. Stat. Solidi B-Basic Solid State Phys. 2006, 243, 3197–3203. (17) Gooding, J. J. Electrochim. Acta 2005, 50, 3049–3060. (18) Wang, J. Electroanalysis 2005, 17, 7–14. (19) Lin, Y.; Taylor, S.; Li, H. P.; Fernando, K. A. S.; Qu, L. W.; Wang, W.; Gu, L. R.; Zhou, B.; Sun, Y. P. J. Mater. Chem. 2004, 14, 527–541. (20) Xu, J. Proc. IEEE 2003, 91, 1819–1829. (21) Avouris, P. Acc. Chem. Res. 2002, 35, 1026–1034. (22) Kamat, P. V. Nano Today 2006, 1, 20–27. (23) Portney, N. G.; Ozkan, M. Anal. Bioanal. Chem. 2006, 384, 620–630. (24) Alvaro, M.; Atienzar, P.; la Cruz, P.; Delgado, J. L.; Troiani, V.; Garcia, H.; Langa, F.; Palkar, A.; Echegoyen, L. J. Am. Chem. Soc. 2006, 128, 6626– 6635. (25) Gruner, G. Anal. Bioanal. Chem. 2006, 384, 322–335. (26) Gong, K. P.; Yan, Y. M.; Zhang, M. N.; Su, L.; Xiong, S. X.; Mao, L. Q. Anal. Sci. 2005, 21, 1383–1393. (27) Khlobystov, A. N.; Porfyrakis, K.; Britz, D. A.; Kanai, M.; Scipioni, R.; Lyapin, S. G.; Wiltshire, J. G.; Ardavan, A.; Nguyen-Manh, D.; Nicholas, R. J.; Pettifor, D. G.; Dennis, T. J. S.; Briggs, G. A. D. Mater. Sci. Technol. 2004, 20, 969–974. (28) Krive, I. V.; Shekhter, R. I.; Jonson, M. Low Temp. Phys. 2006, 32, 887–905. (29) Ye, X.; Gu, X.; Gong, X. G.; Shing, T. K. M.; Liu, Z. F. Carbon 2007, 45, 315–320. (30) Monthioux, M. Carbon 2002, 40, 1809–1823. (31) Naguib, N.; Ye, H. H.; Gogotsi, Y.; Yazicioglu, A. G.; Megaridis, C. M.; Yoshimura, M. Nano Lett. 2004, 4, 2237–2243. (32) Svrcek, V.; Pham-Huu, C.; Amadou, J.; Begin, D.; Ledoux, M. J.; Le Normand, F.; Ersen, O.; Joulie, S. J. Appl. Phys. 2006, 99. (33) Yanagi, K.; Iakoubovskii, K.; Matsui, H.; Matsuzaki, H.; Okamoto, H.; Miyata, Y.; Maniwa, Y.; Kazaoui, S.; Minami, N.; Kataura, H. J. Am. Chem. Soc. 2007, 129, 4992–4997.
Figure 2. Amounts of bound RC protein (in mol/cm2) after its incubation with various substrates (indicated) for 1 h. Error bar ) ( one standard deviation.
fresh surface (basal plane) was created by cleaving a thin layer of carbon using a double-sided Scotch tape. The carbon nanotube array electrode was fabricated by a template-assisted chemical vapor deposition (CVD) growth process.20 To exclude the tube exposure above the spacer surface they were subjected to postgrowth reactive ion etching eating the tubes down to 2-5 nm below the template/spacer surface so that no outer CNT walls were exposed to the surrounding medium.20 The tube diameter and the pattern of tubes inside the template/spacer (Al2O3) were identified with transmission electron microscopy (TEM) and atomic force microscopy (AFM). For scanning electron microscopy (SEM) and AFM experiments, nano tube ends were exposed by etching Al2O3 for about 20-30 nm with 20% HCl in water having 0.1 M CuCl2. Topographic images of the CNT array were recorded at ambient conditions with JEOL SPM-5200 microscope; using Si cantilevers with a force constant of 40 N/m in a tapping mode at frequencies around 300 kHz. The RC containing a polyHis tag at close proximity to primary donor (P+) (at the C-terminal end of M-subunit) was isolated from Rb.sphaeroides strain SMpHis as described elsewhere.11,12 The quality of the RC preparation was confirmed by SDS-PAGE electrophoresis (a single band under nondenaturating conditions and three bands of expected molecular weights after protein denaturation), by the protein absorption spectrum (position of the pigment bands and the protein to pigment absorption ratio, A280/A804 ) 1.2 or less), and by kinetics of photoinduced ET within the protein at room temperature (a single exponential reaction between Qb- and P860+ with rate constant about 0.9 s-1). The 5 nm gold nanoparticles, used for protein visualization inside CNT, were purchased from SPI Supplies (West Chester, PA). The particles were concentrated to ∼4 µM in a Centriplus centrifuge at
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Figure 3. (A) Transmission electron micrograph of a cross section of blank arrayed CNT-alumina membrane without RC protein and (B) with RC protein-nanogold conjugates. (C) and (D) are enlarged selection from (A) and (B), respectively. Arrows point RC-nanogold conjugates. The bars in panels A and B are 200 nm and in panels C and D they are 35 and 45 nm, respectively.
Figure 5. Normalized photocurrent as a function of time for growth (A) and decay (B) of the steady state photocurrent for RC immobilized on HOPG and CNT. The electrode potential was set at 0.05 V vs NHE. Illumination at
