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structural integrity of the LH2 membrane protein. Figure 1b showsa representative spectral image for the patterned AdnLH2 complexes.On each pixel, a full spectrum was recorded, and integrated overthe respective emission band of the LH2 complexes. The exposureof patterns of β-CD SAMs surrounded by a protein resistant PEGSAM14to the solution of AdnLH2, 1 mM AdHEG resulted in theselective assembly of the protein onto the β-CD regions (green) ina ratio of 16:1 as indicated by the averaged emission spectra (insetFigure 1b) from active and passivated areas. A monolayer coveragewas suggested by quantitative spectral images, which revealed onlyminor variations (<5%) in intensity over the patterned area.We performed AFM imaging at low tapping amplitudes to assessthe density of the putative monolayer. Figure 1c shows an AFMheight image of the patterned LH2 complexes; analysis of thesurface indicates a uniform height of ∼6 nm (inset) with referenceto the defect (black region), suggesting monolayer coverage of theprotein. In the AFM image (Figure 1d) ring-shaped LH2 proteinscould be observed, which are attributed to the exposed face of thecomplexes. The height histogram (inset Figure 1c) indicated lessthan 1% multilayer stacked aggregates of proteins in agreementwith the fluorescence images.In an attempt to prepare structures approaching moleculardimensions, NIL was performed using stamps with silicon ridgesas small as 40 nm, 4 µm period. Figure 2a shows an AFM heightimage in liquid of AdnLH2 complexes on a β-CD SAM. The meanfwhm of the lines after processing is 80 ( 5 nm with a height of∼6( 1 nm consistent with the assembly of a monolayer of LH2complexes, Figure 2c. The increase in width, relative to that of theNIL stamp, is attributed to the process of removal of the residuallayer in the imprinting process. Figure 2b shows a fluorescenceimage acquired with a single photon counting avalanche photodiode,with intensity variations of (14% along the lines. The apparentwidth of these structures (Figure 2d) is defined by the opticalresolution of ∼700 nm.We have achieved exquisite spatial control at different lengthscales of functional specifically bound LH2 complexes in a highthroughput manner by exploiting host-guest interactions and NIL.In situ characterization of the formation of these assemblies atmolecular dimensions and the fabrication of mixed protein arraysare the subject of current research.Acknowledgment. This work was supported by NanoNedTMM.7124; NWO-CW (M.J.W.L.; Vidi Vernieuwingsimpuls700.52.423 to J.H.); MESA+ (SRO-NF) (Y.Z.); C.N.H. and J.D.O.acknowledge funding from the BBSRC, U.K.Supporting Information Available: Materials and experimentalprocedures. This Material is available free of charge via the Internet athttp://pubs.acs.org.References(1) Mulder, A.; Onclin, S.; Peter, M.; Hoogenboom, J. P.; Beijleveld, H.; terMaat, J.; Garcia-Parajo, M. F.; Ravoo, B. J.; Huskens, J.; van Hulst, N. F.;Reinhoudt, D. N. Small 2005, 1, 242–253.(2) Ludden, M. J. W.; Reinhoudt, D. N.; Huskens, J. J. Chem. Soc. ReV. 2006,35, 1122–1134.(3) Chou, S. Y.; Krauss, P. R.; Renstrom, P. J. Science 1996, 272, 85–87.(4) Maury, P.; Peter, M.; Crespo-Biel, O.; Ling, X. Y.; Reinhoudt, D. N.;Huskens, J. Nanotechnology 2007, 18, 044007.(5) Hunter, C. N.; Tucker, J. D.; Niederman, R. A. Photochem. Photobiol.2005, 4, 1023–1027.(6) Scheuring, S.; Seguin, J.; Marco, S.; Levy, D.; Breyton, C.; Robert, B.;Rigaud, J. 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Microsc. (Oxford) 2005, 217, 109–116.(14) Kannan, B.; Castelino, K.; Chen, F.; Majumdar, A. Biosens. Bioelectron.2006, 21, 1960–1967.JA802843MFigure 1.(a) Fluorescence titration: nonlabeled LH2 (blue box); nonlabeledLH2, 1 mM AdHEG (open box); AdnLH2, 1 mM AdHEG (green star);reference spectrum of nonlabeled LH2 in solution (red triangle). (b) Falsecolor, fluorescent-spectral image of AdnLH2 patterns (β-CD/PEG), 40 ×40 µm, 64 × 64 pixels; inset shows emission spectra active area (greenbox), passivated (open box). (c) AFM topography in liquid, 150 × 150 nmarea, 256 × 256 pixels, inset shows histogram height distribution. (d) Sectionacross a LH2 complex showing a profile along the dotted line; scale bar,10 nm.Figure 2.(a) AFM topography in liquid of AdnLH2 β-CD/PEG SAM, 10× 10 µm, z-scale 30 nm, and respective cross section (c), fwhm of 80 nm(inset). (b) False color fluorescent image. (d) Cross section of panel b.J. AM. CHEM. SOC.9VOL. 130, NO. 28, 2008 8893C O M M U N I C A T I O N S