Interaction of Tyrosine-, Tryptophan-, and Lysine-Containing

Inorganic Chemistry Laboratory, UniVersity of Oxford, South Parks Road, OX1 3QR Oxford, United Kingdom. ReceiVed: July 30, 2007; In Final Form: Septem...
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J. Phys. Chem. C 2007, 111, 18520-18524

Interaction of Tyrosine-, Tryptophan-, and Lysine-Containing Polypeptides with Single-Wall Carbon Nanotubes and Its Relevance for the Rational Design of Dispersing Agents Christoph G. Salzmann,* Michael A. H. Ward, Robert M. J. Jacobs, Gerard Tobias, and Malcolm L. H. Green Inorganic Chemistry Laboratory, UniVersity of Oxford, South Parks Road, OX1 3QR Oxford, United Kingdom ReceiVed: July 30, 2007; In Final Form: September 26, 2007

The polypeptides poly-L-(Trp,Lys‚HBr) (Trp/Lys‚HBr ) n/m) and poly-L-(Tyr,Lys‚HBr) (Tyr/Lys‚HBr ) n/m) were tested as dispersing agents for single-wall carbon nanotubes (SWCNTs) for n/(n+m) values of 0, 0.2, and 1. Best results were obtained for the two copolymers, illustrating the importance of using amphiphilic dispersing agents. The dispersion effect is 3.5 times higher for the copolymer containing tryptophan, indicating a stronger interaction of the tryptophan residue with the SWCNTs compared to tyrosine. The degrees of debundling of the SWCNTs and the apparent binding of the polypeptides with the SWCNTs are analyzed by atomic force microscopy for the different dispersions. The interactions of the aromatic amino acid residues with the SWCNTs are further probed by using optical absorbance and fluorescence spectroscopy.

Introduction The noncovalent interaction between proteins or polypeptides and single-wall carbon nanotubes (SWCNTs) is of fundamental importance for the use of SWCNTs in biosensing, biocompatibility, or delivery applications.1-4 Furthermore, polypeptides adsorbed onto SWCNTs have also been used for the preparation of SWCNT composite materials.5 For unfunctionalized SWCNTs, the π-π stacking interaction between the aromatic amino acids of the polypeptides and the extended π-electron system of the SWCNTs is considered to be the main driving force for polypeptide adsorption onto SWCNTs.6,7 It was shown that proteins adsorbed onto SWCNTs can retain their enzymatic activities8 and also be stabilized at elevated temperatures and in organic solvents.9 Furthermore, protein materials have been shown to assist the dispersion of SWCNTs in water which is an essential step for most of the SWCNT applications. So far, short amphiphilic peptides,10-12 hydrophobic peptides derived from phage displayed peptide libraries,13 branched anionic and cationic peptide amphiphiles,14 reversible cyclic peptides (RCPs),15 and naturally occurring proteins16-18 have been successfully employed. The importance of the amphiphilicity of the proteins has thereby been conjectured. Here we demonstrate the importance of the amphiphilic character of polypeptides in SWCNT dispersion experiments by testing the ability of the synthetic high molecular weight polypeptides poly-L-(Trp,Lys‚HBr) (Trp/Lys‚HBr ) n/m) and poly-L-(Tyr,Lys‚HBr) (Tyr/Lys‚HBr ) n/m) to disperse and debundle SWCNTs for n/(n+m) ratios of 0, 0.2, and 1 (cf. Figure 1). The adsorptive interactions of the polypeptides with the SWCNTs are analyzed by changes in the electronic interband transitions of the SWCNTs and the fluorescence properties of the adsorbed polypeptides. The topography of the SWCNTs with adsorbed polypeptides is studied by atomic force microscopy on a mica surface. * Author to whom correspondence should be addressed. E-mail: [email protected].

Figure 1. The structures of the random copolymers poly-L-(Tyr,Lys‚ HBr) (a) and poly-L-(Trp,Lys‚HBr) (b).

Experimental Section Chemically unfunctionalized SWCNTs produced by the highpressure CO (HiPco) method19 were supplied by Carbon Nanotechnology, Inc (purified grade, lot number P0323). The polypeptides and sodium dodecyl sulfate (SDS) were purchased from Sigma Aldrich. Solutions of 10.22 mg of poly-L-Lys‚HBr (mol wt 15 000-30 000 g mol-1), 9.11 mg of poly-L-tryptophan (mol wt 15 000-50 000 g mol-1), 7.98 mg of poly-L-tyrosine (mol wt 10 000-40 000 g mol-1), 10.00 mg of poly-L-(Trp,Lys‚HBr) (1:4) (random copolymer, mol wt 20 000-50 000 g mol-1), 9.77 mg of poly-L-(Tyr,Lys‚HBr) (1:4) (random copolymer, mol wt 20 000-50 000 g mol-1), 8.85 mg of polyL-(Trp,Arg‚HCl) (random copolymer, mol wt 20 000-50 000 g mol-1), and 14.1 mg of SDS were prepared by dissolving in 10 mL H2O or D2O. The resulting concentrations of amino acid residues or SDS are 5 mmol L-1. The complete dissolution of poly-L-tryptophan and poly-L-tyrosine seemed difficult, and turbid suspensions were obtained even after stirring overnight. The polypeptide solutions were then combined with 2.50 mg of the SWCNTs, and the SWCNTs were dispersed by ultrasonic agitation in a bath sonicator (110 W) for 30 min. All dispersions were allowed to settle, and the supernatant was passed through densely packed glass wool before analysis.

10.1021/jp076013h CCC: $37.00 © 2007 American Chemical Society Published on Web 11/21/2007

Single-Wall Carbon Nanotubes Raman spectra of the dispersions were recorded as described previously in ref 20 using a Jobin Yvon spectrometer (Labram 1B) equipped with a microscope, through a 10 fold magnification objective (Olympus company), by coadding four spectra with collection times of 40 s each. A 20 mW He-Ne laser (632.8 nm) was used, and the 1800 L/mm grating provides a resolution starting from 1.0 cm-1 at 200 cm-1 up to 0.5 cm-1 at 3600 cm-1. The abscissa was calibrated with the 520.7 cm-1 peak of a silicon standard, and the sharp Raman shifts are accurate within the limits of the resolution. UV-visible-NIR absorbance spectra were recorded in quartz cuvettes (light path length of 10 mm) on a Varian Cary 5000 spectrometer in the range from 1800 to 200 nm with a scan rate of 60 nm min-1 and a resolution of 0.5 nm. A Digital Instruments Multimode SPM atomic force microscope (AFM) was used with a Nanoscope IIIa controller, operating in tapping mode with an ‘E’ scanner, having a lateral range of 12 mm and a vertical range of 3.5 mm. Silicon probes (Nascatec GmbH model NST-NCHFR), with resonant frequencies of approximately 320 kHz were used. X, Y, and Z calibration of the AFM was accomplished by scanning a 10 µm pitch 200 nm 3D reference from Digital instruments. The SWCNT dispersions were dropped onto freshly cleaved mica surfaces using a Laurell Technologies WS-400 spin-coater (3000 rpm). Fluorescence spectra were recorded in quartz cuvettes (light path length of 10 mm) on an F-4500 FL spectrophotometer with excitation wavelengths from 200 to 600 nm in steps of 5 nm. The emission spectra were collected between 200 and 800 nm with a scan rate of 1200 nm min-1 and a resolution of 5 nm. Results and Discussions The Raman spectroscopic analysis of the SWCNT dispersions is shown in Figure 2. The technique used here for quantitative analysis is described in detail in ref 20 and allows a semiquantitative determination of the SWCNT concentration in dispersions. Thereby, the areas of the SWCNT G-bands centered at 1589 cm-1 are normalized with respect to the νO-H peak of water at ∼3300 cm-1 and compared to the G-band area of a benchmark dispersion. Here, the SDS dispersion (spectrum (7)) serves as the benchmark and the G-band areas of the dispersions using the polypeptide solutions (spectra (2-6)) are given as percentages of the G-band area obtained for the SDS dispersion (%SDS). SDS is currently one of the most effective dispersing agents for SWCNTs.20,21 For pure water (spectrum (1)), the G-band area is, as expected, 0.0 %SDS. The dispersion using poly-L-Lys‚HBr shows a small dispersion effect of 2.9 %SDS (spectrum (2)). The G-band areas increase to 6.9 %SDS (spectrum (3)) and 23.9 %SDS (spectrum (4)) for poly-L-(Tyr,Lys‚HBr) and poly-L-(Trp,Lys‚HBr), respectively. For the homopolymers of tyrosine (spectrum (5)) and tryptophan (spectrum (6)), the G-band areas decrease to 0.3 %SDS and 0.0 %SDS, respectively. A plot of the obtained G-band areas for the different dispersion experiments as a function of the content of aromatic amino acids (n/(n+m)) in the polypeptides is shown as inset in Figure 2. These results illustrate two prerequisites for using polypeptides as dispersing agents: First, a polypeptide has to have affinity to adsorb onto the surface of SWCNTs. As stated previously, the most important force for noncovalent adsorption is the π-stacking of aromatic amino acids such as tyrosine or tryptophan with the extended π-system of the SWCNTs. Second, in order to achieve high degrees of dispersion, the adsorbed polypeptide has to exhibit hydrophilic properties originating

J. Phys. Chem. C, Vol. 111, No. 50, 2007 18521

Figure 2. Raman spectra of the SWCNT dispersions normalized with respect to the area of the νO-H peak (λ0 ) 632.8 nm) (cf. ref 20). The dispersions were prepared using (1) pure water and solutions of (2) poly-L-Lys‚HBr, (3) poly-L-(Tyr,Lys‚HBr), (4) poly-L-(Trp,Lys‚HBr), (5) poly-L-Tyr, (6) poly-L-Trp, and (7) sodium dodecyl sulfate (SDS). The areas of the G-bands are given as percentages of the G-band area of the SDS dispersion (7). The peaks arising from SWCNTs (D, G, and G′ bands) and water (νO-H) are indicated. (Inset) Plot of the G-band areas (as percentages of the value obtained for the SDS dispersion) as a function of the content of tryptophan (solid squares) or tyrosine (solid diamonds) in the various polypeptides (n/(n+m)) (cf. Figure 1). Dashed lines are guides to the eye.

from hydrophilic amino acid residues such as lysine hydrobromide in order to make the SWCNT/polypeptide conjugate soluble. Interestingly, we find here that poly-L-Lys‚HBr shows a small dispersing effect despite its lack of aromatic amino acid residues. In general, charged particles or ions can be used to stabilize dispersions. For SWCNTs, this was previously demonstrated by using charged zirconia nanoparticles,22 and it is assumed that the same effect could play a role here in the dispersion experiment using poly-L-Lys‚HBr. Due to the positive charges on the side chain of the lysine residues, the polypeptide carries a high density of positive charge. This positively charged polymer haloing22 or perhaps even partially wrapping the SWCNTs could be responsible for stabilizing the dispersion through an electrostatic effect and thereby preventing the SWCNTs from reaggregating. However, the dispersion effect observed for poly-L-Lys‚HBr (2.9 %SDS) is comparatively small compared to the dispersions using poly(Tyr,Lys‚HBr) (spectrum (3), 6.9 %SDS) and especially poly(Trp,Lys‚HBr) (spectrum (4), 23.9 %SDS). The aromatic amino acids in these copolymers provide an additional mechanism for adsorption onto the surface of the SWCNTs. The increased amounts of adsorbed polypeptide stabilize the dispersion to a greater extent by increasing the electrostatic repulsion between the different polypeptide/ SWCNT conjugates. The higher dispersion effect observed for poly(Trp,Lys‚HBr) compared to poly(Tyr,Lys‚HBr) indicates a stronger π-interaction of the tryptophan residues with the SWCNTs compared to the tyrosine residues. This could be explained by the larger π-system of the tryptophan residue (cf. Figure 1) allowing a more pronounced π-stacking effect. Additionally, tryptophan has a higher hydrophobicity index (1.9) than tyrosine (-0.7)23 which creates a more hydrophobic environment in the interior of the poly-(Trp,Lys‚HBr) where especially the longer sequences of the aromatic amino acids are expected to reside. The

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Figure 3. Atomic force microscopy images of the SWCNT dispersions deposited onto mica. The dispersing agents are poly-L-Lys‚HBr (a), poly-L-(Trp,Lys‚HBr) (b,d), and poly-L-(Tyr,Lys‚HBr) (c). The arrows indicate adsorbed polypeptide in b and c, and end-to-end linking of SWCNTs in d. Z-scales are given for all figures.

importance of tryptophan residues for the interaction of polypeptides or proteins with SWCNTs was also previously shown by a large increase of the fraction of tryptophan residues after a phage display experiment.13 The homopolymers of the aromatic amino acids (poly-Tyr and poly-Trp) show a high affinity toward the SWCNTs as the originally turbid polypeptide solutions become clear after ultrasonication with the SWCNTs and settling of the SWCNTs. The lack of charge on the adsorbed homopolymers is probably the reason for the small dispersion effect of 0.3 %SDS for polyTyr (spectrum (5)) and 0.0 %SDS for poly-Trp (spectrum (6)). In fact, the adsorbed homopolymers of tyrosine and tryptophan might even increase the affinity of the SWCNTs to re-aggregate through π-stacking interactions between the adsorbed polypeptides. The Raman spectrum of the SWCNT dispersion in SDS solution shows a broad underlying peak (spectrum (7)) which is thought to arise from the presence of fluorescing carbonaceous compounds in the dispersion.20,24,25 For the polypeptide dispersions (spectra (2-6)), this fluorescence peak is not observed which indicates that the polypeptides disperse SWCNTs more specifically than SDS. This specificity toward SWCNTs is attributed to the polymeric nature of the polypeptides which seems to favor adsorption on the large length scale SWCNTs. The abilities of the polypeptides to debundle SWCNTs were analyzed next by atomic force microscopy (AFM). From AFM analysis of the poly-L-Lys‚HBr dispersion (cf. Figure 3(a)), it can be seen that poly-L-Lys‚HBr does not completely debundle the SWCNTs. For this sample, no individual SWCNTs could be found on the mica surface. In fact, the polypeptide seems only to be interwoven with larger SWCNT aggregates. Also, the strands of single SWCNTs showing out of the agglomerate are not wrapped by the polypeptide. For the poly-L-(Tyr,Lys‚HBr) and the poly-L-(Trp,Lys‚HBr) dispersions, however, only monodispersed SWCNTs or very small bundles were observed in AFM measurements (cf. Figure 3(b,c)). This shows the importance of the aromatic amino acid

Salzmann et al. residues in the debundling process and the efficiency of the charged copolymers to stabilize the (mono)dispersions. Because of the large difference in the hydrophobicity indices of lysine (-8.8) and tyrosine (-0.7) or tryptophan (1.9),26 a globular shape of the polypeptides is expected in solution, with longer sequences of lysine hydrobromide more likely to be found on the outside of the polypeptides thereby maximizing the distances between the positive charges on the lysine side chains, and longer sequences of tyrosine or tryptophan buried in the hydrophobic interiors of the polypeptides. The globular shape of the polypeptides seems to be retained to some extent after the adsorption of the polypeptides onto the SWCNTs (cf. Figure 3(b,c)). Generally, a “beads-on-a-string” binding geometry is observed for both copolymers. The volume of a polypeptide can be estimated by using the generally accepted value of ∼1.37 g cm-3 as density.27 From the molecular weights of the copolymers, this gives a molecular volume of 42 ( 18 nm3. For spherically shaped polypeptides, this corresponds to a diameter of 4.3 ( 0.7 nm. AFM height analysis of the beads indicated by arrows in Figure 3(b,c) gives an average height of 2.3 ( 0.4 nm for adsorbed poly-L-(Trp,Lys‚HBr) and 2.2 ( 0.2 nm for adsorbed poly-L-(Tyr,Lys‚HBr). Considering that these heights include the incorporated SWCNTs and that the density of the polypeptides is probably slightly increased due to the loss of water, this seems to indicate that the beads found on the SWCNTs consist of single polypeptide molecules separated from each other by the electrostatic repulsion of the charged lysine residues. Unbound polypeptide can be seen as also spherical objects on the AFM substrate in Figure 3(b-d). It seems likely that the globular copolymers will unfold to some extent during the ultrasonication process and incorporate the SWCNTs into their hydrophobic interiors. The high affinity of the poly-L-(Trp,Lys‚HBr) copolymer with the SWCNTs is furthermore illustrated by the AFM observation of the polypeptide incorporating the ends of two SWCNTs and linking them together. The area shown in Figure 3(d) shows two spots where two SWCNTs are linked together end-to-end by the polypeptide (marked by arrows). A similar linking behavior was previously reported for SWCNTs wrapped with the nano1 peptide which could potentially allow the assembly of SWCNTs into macromolecular structures.28 In AFM analysis of the dispersion using the homopolymers of tyrosine and tryptophan, consistent with the results from the Raman spectroscopic analysis (cf. Figure 2), no dispersed SWCNTs could be found. Instead rather large agglomerates of the polypeptides were identified on the AFM substrates (not shown). The effects of SWCNT debundling can also be seen from the optical absorbance measurements shown in Figure 4. In the UV-vis-NIR spectral range, the electronic interband transitions of the SWCNTs are observed.29,30 These arise from transitions between the sharp van Hove maxima in the electronic density of states of the SWCNTs. The transition energies depend mainly on the tube diameters.31 For HiPco SWCNTs, the S11 and the S22 transitions are typically found in the ranges from 800 to 1600 nm and 550 to 900 nm, respectively.30 The broadness of the observed features in optical absorbance spectroscopy is thereby indicative of the degree of debundling.30 The peak at ∼1105 nm (marked by arrow) located on the shoulder of the main peak at ∼1200 nm becomes increasingly less resolved in the sequence SDS, poly-L-(Trp,Lys‚HBr), poly-L-(Tyr,Lys‚HBr), and poly-L-Lys‚HBr which indicates a decrease of the fraction of monodispersed SWCNTs in the same order.

Single-Wall Carbon Nanotubes

Figure 4. Optical absorbance spectra of the SWCNT dispersions in solutions of sodium dodecylsulfate (black), poly-L-(Trp,Lys‚HBr) (red), poly-L-(Tyr,Lys‚HBr) (green), and poly-L-Lys‚HBr (blue). The spectra are normalized for the intensity of the peak at ∼1200 nm and shifted vertically for clarity. All dispersions were measured in D2O solutions. Peaks marked with asterisks arise from small H2O/HDO impurities. (Inset) Magnification of the normalized absorbance spectra around ∼1200 nm (axes have the same units as the main figure). Spectra are not vertically shifted.

The peaks marked with asterisks in Figure 4 originate from small impurities of H2O or more likely HDO in the D2O solutions. This was shown by adding a small amount of H2O to one of the dispersions which lead to an increase in the intensities of the peaks in question. Also, solutions of just the polypeptides in D2O showed these peaks (not shown). This indicates that the received polypeptide samples contain small amounts of H2O. The peak position of the main peak centered around 1200 nm increases from 1179 nm, to 1187, 1190, and 1193 nm for the SDS, the poly-L-Lys‚HBr, the poly-L-(Tyr,Lys‚HBr), and the poly-L-(Trp,Lys‚HBr) dispersion, respectively (cf. inset in Figure 4). It is well-known that the SWCNTs in SDS, as well as sodium dodecylbenzenesulfonate (SDBS) dispersions, show significantly blue-shifted peaks in optical absorbance spectroscopy. For most other dispersing agents, considerably more redshifted peaks are observed.21 This is also what we find here for the poly-L-Lys‚HBr, the poly-L-(Tyr,Lys‚HBr), and the polyL-(Trp,Lys‚HBr) dispersion. It seems as if the increasingly stronger π-interaction of the polypeptides with the SWCNTs leads to a red-shift of the peaks. This trend is also found for the other peaks in the S11 and S22 spectral ranges. The adsorptive interactions of poly-L-(Trp,Lys‚HBr) and polyL-(Tyr,Lys‚HBr) with the SWCNTs have also been probed by fluorescence spectroscopy. Both, tryptophan and tyrosine amino acid residues are well-known to exhibit fluorescence behavior.32,33 First, the dispersion of SWCNTs in poly(Trp,Lys‚HBr) solution was analyzed (cf. Figure 5(a)). Apart from the fluorescence peak of the tryptophan residues, a strong Rayleigh scattering line is observed, arising from the dispersed SWCNTs. The dispersion was then filtered three times through the same Teflon membrane (0.2 µm pore diameters) in order to remove the SWCNT/adsorbed polypeptide conjugates. The successful removal can be seen from the almost complete disappearance of the Rayleigh scattering line (cf. Figure

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Figure 5. Contour plots (linear color gradient) of the fluorescence spectra of (a) the SWCNT dispersion in poly-L-(Trp,Lys‚HBr) solution, (b) the filtrate of the dispersion in poly-L-(Trp,Lys‚HBr) solution, (c) the difference spectrum between a and b, (d) the SWCNT dispersion in poly-L-(Tyr,Lys‚HBr) solution, (e) the filtrate of the dispersion in poly-L-(Tyr,Lys‚HBr) solution, and (f) the difference spectrum between d and e. Spectra a, b, d, and e are normalized with respect to the maxima of the fluorescence peaks. Spectra c and f are shown on a 10 times smaller scale than a, b, d, and e. The arrow in c indicates the shift of the peak maximum from b to c.

5(b)) and was also confirmed by Raman spectroscopy and optical absorbance spectroscopy (not shown). The original dispersion contained SWCNTs with adsorbed polypeptide and free polypeptide. After filtration, only the free polypeptide remained in solution. By subtracting spectra b from a, the fluorescence spectra of the SWCNT/adsorbed polypeptide conjugate were obtained (cf. Figure 5(c)). It can be seen that both the excitation wavelength and to a smaller degree the emission wavelength of the peak maximum, experience a redshift (indicated by arrow). A similar red-shift in the emission spectrum has also been observed for anthracene derivatives adsorbed onto SWCNTs.34 (The effect on the excitation wavelength was not tested in ref 34.) We interpret these redshifts as consequences of the adsorptive interaction of the tryptophan amino acid residues with the SWCNTs through π-stacking. This fluorescence spectroscopic analysis was carried out next for the SWCNT dispersion in the poly(Tyr,Lys‚HBr) solution (cf. Figure 5(d-f)). In this case, the adsorbed poly(Tyr,Lys‚ HBr) shows a smaller fluorescence intensity (cf. Figure 5(f)) which is, consistent with the finding above, probably due to the lower concentration of strongly adsorbed tyrosine residues. Generally, the fluorescence peak seems to broaden only for the adsorbed polypeptide, predominately in the direction of the excitation wavelength. This indicates a smaller degree of adsorptive interaction between poly-L-(Tyr,Lys‚HBr) and the SWCNTs compared to the poly-L-(Trp,Lys‚HBr) interaction which is consistent with the dispersing effects of the two copolymers found above.

18524 J. Phys. Chem. C, Vol. 111, No. 50, 2007 Conclusions We have illustrated the importance of the balance between (charged) hydrophilic and aromatic amino acid residues in polypeptides for the successful dispersion of SWCNTs. In this context, tryptophan amino acid residues interact more strongly with the SWCNTs than tyrosine residues making the copolymer of tryptophan and lysine hydrobromide a better dispersing agent for SWCNTs compared to the copolymer of tyrosine and lysine hydrobromide. Both copolymers are capable of debundling SWCNTs and also stabilizing the dispersion for at least several months. In order to investigate the influence of the nature of the hydrophilic amino acids residues, poly-L-(Trp,Arg‚HCl) was additionally tested as a dispersing agent which increased the value of the normalized G-band area by only 1.7 %SDS compared to the value obtain for poly-L-(Trp,Lys‚HBr) (not shown). With a hydrophobicity index of -12.3, arginine is the most hydrophilic of the naturally occurring amino acid residues.23 For comparison, the hydrophobicity index of lysine is -8.8.23 This rather small increase of G-band area indicates that the nature of charged hydrophilic part of the copolymers is not the limiting factor in achieving higher degrees of dispersion. We believe that the optimization of the n/(n+m) ratio and/or the use of more hydrophobic, perhaps synthetic amino acid residues with large π-systems could lead to high dispersing properties at ideally small n/(n+m) values. Acknowledgment. We acknowledge the Austrian Academy of Sciences (Austrian Program for Advanced Research and Technology) (C.G.S.), the EPSCR (M.A.H.W.), and the EC Marie Curie Intra-European Fellowship Program (MEIF-CT2006-024542) (G.T.) for financial support. Furthermore, we thank Begbroke Nano for access to the UV-vis-NIR spectrometer, and Alison Crossley and Colin Johnston for helpful discussions. References and Notes (1) Bianco, A.; Prato, M. AdV. Mater. 2003, 15, 1765. (2) Lin, Y.; Taylor, S.; Li, H.; Shiral, Fernando, K. A.; Qu, L.; Wang, W.; Gu, L.; Zhou, B.; Sun, Y.-P. J. Mater. Chem. 2004, 14, 527. (3) in het Panhuis, M.; Gowrisanker, S.; Vanesko, D. J.; Mire, C. A.; Jia, H.; Xie, H.; Baughman, R. H.; Musselman, I. H.; Gnade, B. E.; Dieckmann, G. R.; Draper, R. K. Small 2005, 1, 820. (4) Kane, R. S.; Stroock, A. D. Biotechnol. Prog. 2007, 23, 316. (5) Pender, M. J.; Sowards, L. A.; Hartgerink, J. D.; Stone, M. O.; Naik, R. R. Nano Lett. 2006, 6, 40. (6) Zorbas, V. Z.; Smith, A. L.; Xie, H.; Ortiz-Acevedo, A.; Dalton, A. B.; Dieckmann, G. R.; Draper, R. K.; Baughman, R. H.; Musselman, I. H. J. Am. Chem. Soc. 2005, 127, 12323.

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