Layer-by-Layer Self-Assembly of Polymer Films and Capsules through

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Layer-by-Layer Self-Assembly of Polymer Films and Capsules through Coiled-Coil Peptides Adam J. Gormley,† Rona Chandrawati,† Andrew J. Christofferson,‡ Colleen Loynachan,† Coline Jumeaux,† Arbel Artzy-Schnirman,† Daniel Aili,§ Irene Yarovsky,‡ and Molly M. Stevens*,† †

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Department of Materials, Department of Bioengineering, and Institute of Biomedical Engineering, Imperial College London, London SW7 2AZ, United Kingdom ‡ Health Innovations Research Institute and School of Aerospace, Mechanical and Manufacturing Engineering, RMIT University, Melbourne, Victoria 3001, Australia § Division of Molecular Physics, Department of Physics, Chemistry and Biology (IFM), Linköping University, Linköping SE-581 83, Sweden S Supporting Information *

ABSTRACT: The layer-by-layer (LbL) technique is a simple and robust process for fabricating functional multilayer thin films. Here, we report the use of de novo designed polypeptides that self-assemble into coiled-coil structures (four-helix bundles) as a driving force for specific multilayer assembly. These pH- (sensitive between pH 4 and 7) and enzymeresponsive polypeptides were conjugated to polymers, and the LbL assembly of the polymer−peptide conjugates allowed the deposition of up to four polymer−peptide layers on planar surfaces and colloidal substrates. Stable hollow capsules were obtained, and by taking advantage of the peptide’s susceptibility to specific enzymatic cleavage, release of encapsulated cargo within the carriers can be triggered within 2 h in the presence of matrix metalloproteinase-7. The enormous diversity of materials that can form highly controllable and programmable coiled-coil interactions creates new opportunities and allows further exploration of the multilayer assembly and the formation of carrier capsules with unique functional properties.

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that can self-assemble into coiled-coil structures (Figure 1). De novo designed polypeptides that self-assemble into coiled-coils

elf-assembly has emerged as a promising and practical way for the creation of materials with great control over structure and functionality. In particular, the layer-by-layer (LbL) technique has proven as a simple and inexpensive process with incredible versatility in generating stable multilayer films and colloidal capsules with precise control over film properties. Principally, a general LbL deposition is achieved using oppositely charged materials (polymers, proteins, nanoparticles, etc.) that deposit through electrostatic interactions.1,2 Such polyelectrolyte multilayer thin films can harbor a wide range of physicochemical properties that allows for precise tailoring of film functions and performances. However, relying on electrostatics to build the multilayer films limits the tailorability of the assembly/disassembly process. To address this limitation, other interactions have been employed to regulate the bonding between the multilayers. Covalent bonding, for example, has been described to generate more robust films.3 On the other hand, hydrogen bonding has been used to incorporate pH- and temperature-responsive characteristics in the multilayer films.4−6 Other molecular interactions such as DNA hybridization7−9 and streptavidin−biotin binding10,11 have also been used to direct the LbL assembly process. These interactions are particularly interesting as their design can dictate the specificity of the multilayers. This is particularly relevant when stimuli-responsive behavior is desired. This work seeks to add a further specific molecular tool to the growing repertoire of multilayer assembly, namely peptides © 2015 American Chemical Society

Figure 1. Schematic illustration of LbL self-assembly through coiledcoil peptides on (a) planar surfaces and (b) colloidal substrates. Alternating polymer−peptide conjugates (i and ii) are deposited onto the surfaces to form multilayer films where self-assembly of the polypeptides into four-helix bundle coiled-coil structures is used as the specific molecular interaction. Upon dissolution of the silica templates (iii), stable hollow capsules can be obtained. Received: July 6, 2015 Revised: August 4, 2015 Published: August 13, 2015 5820

DOI: 10.1021/acs.chemmater.5b02514 Chem. Mater. 2015, 27, 5820−5824

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Chemistry of Materials

the loop region of the peptides via thiol-maleimide chemistry. We chose 42-residue polypeptides JR2EC and JR2KC (helix− loop−helix motifs) previously used in our lab that are known to self-assemble into four-helix bundles in solution, as shown by circular dichroism (CD) spectroscopy (Figure 2b−d).14,19,20 These peptides comprise a very robust and convenient system for specific folding-dependent assembly of nanomaterials.19,21 They also exhibit a large extent of tunability, which has been utilized for realizing a number of different enzyme-responsive materials for bioanalytical applications.14,22 The resulting conjugates contain six peptides per polymer chain as confirmed by NMR. CD analysis indicates that polymer attachment provided slightly increased ellipticity due to entropic stabilization, though not enough to form coiled-coils until mixed together (Figure S1). The ability of these peptides to form coiled-coil cross-links while attached to the copolymers was confirmed by dissolving both polymer−peptide conjugates individually in PBS (10 mg/mL) followed by mixing of the two solutions. As expected, this resulted in the formation of a polymer−peptide hydrogel.13 Quartz crystal microbalance (QCM) was used to monitor the buildup of the multilayer assembly of the polymer−peptide conjugates on planar surfaces (Figure 3a). First, a JR2EC

are uniquely suited for biomedical applications as the peptide sequences, and therefore structures can be used to tailor the binding strengths of the multilayer films and the stimuliresponsive characteristics.12 For example, coiled-coils have been used to cross-link hydrogels that swell in response to changes in pH.13 We recently demonstrated that coiled-coil peptides can specifically regulate nanoparticle assembly for the development of bioassays.14,15 Finally, coiled-coils have been used for surface functionalization and shown to exhibit stimuli-responsive characteristics.16−18 These examples highlight the immense utility of coiled-coils in the design and preparation of responsive, self-assembled functional materials. In this study, we: (i) synthesize polymer−peptide conjugates as the building blocks for the LbL assembly; (ii) demonstrate the formation of multilayer films via coiled-coil interactions on planar surfaces and colloidal substrates; (iii) model the polymer−peptide films and show pH-responsive characteristics; (iv) characterize the structural integrity of the polymer−peptide carrier capsules; and (v) investigate the stimuli-responsive characteristics of the multilayer films in the presence of matrix metalloproteinase-7 (MMP-7). To demonstrate the formation of multilayer LbL films via coiled-coil interactions, the polypeptides were conjugated to a N-(2-hydroxypropyl)methacrylamide-co-N-(3-aminopropyl)methacrylamide (HPMA-co-APMA) copolymer backbone. Copolymers of HPMA (90 mol %) and APMA (10 mol %) were first synthesized via reversible addition−fragmentation chain transfer (RAFT) polymerization (Figure 2a). The

Figure 3. (a) QCM frequency shifts upon deposition of multilayer of polymer−peptide conjugates (1−4) on planar surfaces. A decrease in frequency indicates an increase of mass adsorbed onto the surface. (b) Stimulated emission depletion (STED) and (c) scanning electron microscopy (SEM) images of hollow capsules obtained via the LbL assembly of four polymer−peptide layers. For imaging, fluorescently labeled HPMA-JR2EC was used.

peptide solution (100 μM in PBS) was introduced onto the QCM chip and allowed to decorate the surface via gold− thiolate bond to the cysteine residues in the center of the peptides to form a precursor peptide layer. It is important to note here that JR2KC would not be a suitable precursor peptide layer as its lysine residues have been shown to bind to the gold surface and prevent proper display of the peptide.23 After being washed with PBS, the chip was exposed to a solution of HPMA-JR2KC (1 mg/mL in PBS) and allowed to incubate for 15 min. A significant decrease in frequency confirms the interaction of HPMA-JR2KC (Figure 3a, 1) with the precursor JR2EC layer via coiled-coils, and the film remained stable after the washing step. Subsequent alternating adsorption of the polymer−peptide conjugates (Figure 3a, 2, HPMA-JR2EC; 3, HPMA-JR2KC; 4, HPMA-JR2EC) can be followed by the constant decrease in frequency, indicating an increase of mass adsorbed onto the surface. It was found that a maximum of four polymer−peptide layers can be deposited,

Figure 2. (a) Chemical structure of polymer−peptide conjugates. Copolymers of HPMA and APMA were synthesized via RAFT polymerization. (b, c) Polypeptides (JR2EC or JR2KC), which form four-helix bundle coiled-coil structures, were conjugated to the polymer backbone via a bifunctional linker SMCC. The polymer− peptide conjugates were fluorescently labeled with fluorescein Omethacrylate (FMA). (d) CD spectra of polypeptides JR2EC and JR2KC confirming the presence of a coiled-coil structure.

resulting precursor copolymers were 18 kDa in molecular weight with narrow dispersities (Đ = 1.08), as determined by size-exclusion chromatography (SEC). To provide a site for peptide attachment, a bifunctional linker 4-(Nmaleimidomethyl)cyclohexanecarboxylic acid N-hydroxysuccinimide ester (SMCC) was coupled to the polymer’s primary amines, and such functionalization allowed for covalent attachment to the cysteine residue in position 22 located in 5821

DOI: 10.1021/acs.chemmater.5b02514 Chem. Mater. 2015, 27, 5820−5824

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Chemistry of Materials

charged residues are more buried, while the peptide surface is dominated by the positively charged lysines. It can be suggested that this is due to the excess of positively charged residues in the peptides, with 16 lysines and two arginines, compared to four aspartic acids and eight glutamic acids, for each dimer. In the relatively constrained polymer−peptide conjugate system, the negatively charged residues interact with the positively charged residues of the adjacent peptide dimer, while the surplus lysines minimize the interpeptide electrostatic repulsion by extending into solution where they interact with dissolved anions (Figure 4a, Figure S10). This in effect facilitates stacking of the dimers within the film.

and after that, no further adsorption was observed likely due to the decreasing density of unbound peptides and therefore a decrease of polyvalent binding. The total decrease in frequency for the LbL assembly of four polymer−peptide layers via coiledcoil interaction was determined to be ∼300 Hz, which is an order of magnitude higher than those obtained by conventional LbL systems (e.g., via electrostatic interaction24 or hydrogen bonding25), suggesting the formation of a thick film. In a control experiment, layering of the peptides without the polymer backbone shows that the multivalent nature of the polymer−peptide conjugates is responsible for the LbL assembly (Figure S2). On the basis of our findings on planar surfaces, we performed the sequential deposition of polymer−peptide conjugates on silica particle templates. Mesoporous silica particles were prepared and were 1.6 μm in diameter (Figure S3). These particles were functionalized with 3-aminopropyltriethoxysilane (APTES) and then SMCC to provide a maleimide group on the particle surface. Subsequent attachment of JR2KC via thiolmaleimide chemistry resulted in particles with a peptide anchor, which was followed by alternating deposition of HPMA-JR2EC and HPMA-JR2KC layers (2 mg/mL in PBS, 15 min). Upon dissolution of the silica particle templates, stable hollow capsules were obtained as confirmed by fluorescence microscopy (Figure S4). For fluorescence microscopy imaging, fluorescently labeled HPMA-JR2EC conjugates were used in the LbL assembly, prepared by copolymerization in the presence of 2 mol % fluorescein O-methacrylate (FMA) (Figure 2a). A stimulated emission depletion (STED) microscopy image of the fluorescently labeled hollow capsule is shown in Figure 3, panel b, confirming the structural integrity. Removal of the sacrificial templates resulted in a slight decrease in the capsule diameter to ∼1.3 μm. Scanning electron microscopy (SEM) images showed the dried capsule appeared as a deflated balloon (Figure 3c, Figure S5), which has previously been observed for hollow capsules formed by other LbL techniques.26 To better understand the structure of the coiled-coils and the resulting films, the four-helix bundles were modeled using allatom molecular dynamics simulations (see Supporting Information). It was important to model this system to (a) visualize the folding of the four-helix bundles, which has not previously been done with these peptides, and (b) understand how the layers assembled in solution relative to one another and explain some unexpected findings in the experiments. The simulated JR2KC/JR2EC dimer produced two different stable structures (Models 1 and 2, Figure S7). Model 1 is in accord with the structure previously proposed by Rydberg and colleagues,20 but Model 2, while slightly offset, was found to be energetically more favorable. This may help to explain the slightly offset ellipticity in the CD spectra at 208 and 222 nm. Unfortunately, a crystal structure of the dimer could not be obtained to confirm the modeling results. The polymer−peptide conjugates were modeled wherein six individual polymer chains were included in the simulation, with three peptides per chain, for a total of nine JR2EC/JR2KC peptide dimers present. In contrast to the isolated peptide dimers, the electrostatic potential maps for the polymer− peptide conjugates show a much greater positive charge concentration at the peptide surface, with little difference between the systems at pH 4 and pH 7. While in both cases there is still a pattern of alternating positive and negative charges, in the polymer−peptide systems, the negatively

Figure 4. Model structure of the JR2EC/JR2KC polymer−peptide conjugates. (a) Interaction between positively and negatively charged residues of adjacent peptide dimers. (b) Model structure at pH 7 shows stacking of the dimers within the film. (c) At pH 4, repulsion of the positively charged dimers results in slight dissociation. JR2EC is colored red, JR2KC is blue, polymers are gray, and negative ions are orange.

The development of therapeutic carriers that can respond to pH changes is of interest for biological systems. We also used the modeling tools to study the changes in the multilayer films in response to pH. As the pH is lowered from pH 7 (assembly condition) to pH 4, it is shown that protonation of the acid residues of the peptides leads to repulsion of the positively charged dimers and therefore results in a slight dissociation of the stacking (Figure 4b,c). These simulation results help to explain the observed changes in the characteristics of the polymer−peptide films upon changes of pH (Figure S11, Figure S12, and Table S1). While in the free peptide dimers denaturation at elevated temperature occurred via an initial dissociation between the helices of the JR2KC peptide, in the polymer−peptide conjugates, the peptides are too tightly packed along the axis of the polymer to allow for this to occur. It can, therefore, be suggested that increasing the ratio of HPMA to APMA-SMCC monomers may facilitate this dissociation and improve the sensitivity of the conjugates to pH and temperature. In addition, replacing the surplus lysines in JR2KC (Lys 6, 10, 13, 17, 28, 32, and 39) with small neutral amino acids such as glycine or alanine could provide a less positively charged environment that will enable a more efficient response of the conjugate systems to changes in pH. 5822

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assembly/disassembly process. Naturally derived or de novo designed coiled-coils are enormously diverse, and a wide range of structures are available. The lab of Derek Woolfson has been actively engaged in classifying these structures and has proposed a periodic table of coiled-coils.28 This classification system is very interesting and may be of tremendous value for the development and design of stimuli-responsive nanomaterials where coiled-coils are used as cross-links. The enormous diversity in sequence, design, and structure of coiled-coils provides many exciting opportunities for exploration of systems similar to the one described in this study. Further investigation into how polymers help order and stabilize these coiled-coil cross-links will likely enable new avenues for building functional multilayers.

To demonstrate the application of the polymer−peptide capsules as therapeutic carriers, we investigated the stimuliresponsive characteristics of the multilayer films in the presence of enzymes. It is known that JR2EC is susceptible to enzymatic cleavage by MMP-7,27 which is implicated in many disease sites such as the tumor microenvironment and cardiovascular disease. HPMA-JR2EC was incubated with MMP-7, and detection of the peptide fragments by liquid chromatography mass spectrometry (LCMS) confirmed the hydrolysis of the peptides due to the activity of MMP-7 (Figure S13). We first monitored the enzyme-responsive characteristics of the multilayer films on planar surfaces. QCM chips with deposited polymer−peptide multilayers were incubated with and without MMP-7 at 37 °C, and the resulting shifts in frequency were monitored. As shown in Figure 5, panel a, the presence and



ASSOCIATED CONTENT

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S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.chemmater.5b02514. Detailed experimental procedures and methods as well as supporting figures (PDF)



AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Phone: +44 (0)20 7594 6804. Author Contributions

The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. Notes

The authors declare no competing financial interest.



Figure 5. Stimuli-responsive characteristics of the polymer−peptide multilayer films in the presence of MMP-7. (a) QCM frequency of the multilayer films in the presence of MMP-7 added at 15 min. An increase in frequency change suggests an erosion of the film due to the enzymatic cleavage. (b) MMP-7 can be used to release encapsulated cargo (FITC-dextran) from the polymer−peptide carrier capsules. A decrease in fluorescence intensity of the capsules was observed after incubation with MMP-7 for 2 h as monitored by flow cytometry.

ACKNOWLEDGMENTS A.J.G. acknowledges support of a Whitaker International Scholarship, and M.M.S. acknowledges funding from EPSRC (EP/K020641/1). I.Y. and M.M.S. acknowledge the Australian Research Council Discovery grant (DP140101888) and a generous allocation of resources on the Australian high performance computational infrastructure facilities, NCI, VLSCI, VPAC, and iVEC. D.A. acknowledges the support from the Foundation for Strategic Research (SSF) and the Swedish Research Council (VR). Special thanks to Ciro Chiappini for help with the STED imaging and Shweta Agarwal for help with the SEM imaging.

activity of MMP-7 resulted in an increase of frequency change, suggesting film erosion due to cleavage of JR2EC within the coiled-coil cross-links. We then demonstrated the MMP-7responsiveness on the colloidal substrates. For this study, FITC-dextran (10 kDa), a common model cargo for following macromolecular drug release in LbL systems, was encapsulated within the polymer−peptide capsules by incubating it with the mesoporous silica particle template before layering and core dissolution. Encapsulated dextran was seen within the capsule cavity and polymer shell. Incubation with MMP-7 resulted in destabilization of the capsules and release of the encapsulated cargo after 2 h as monitored by flow cytometry (Figure 5b). In the absence of MMP-7, the capsules remained stable for up to 6 h, and there was negligible release of the fluorescent cargo from the capsules (Figure S14). These results highlight that these capsules, stabilized through JR2EC/JR2KC four-helix bundle coiled-coil structures, have the potential to release therapeutics (e.g., drugs, proteins, nanoparticles) upon exposure to MMP-7. Use of coiled-coils to assemble multilayer films is an attractive technique as it allows for precise tailorability of the



REFERENCES

(1) Hammond, P. T. Recent explorations in electrostatic multilayer thin film assembly. Curr. Opin. Colloid Interface Sci. 1999, 4, 430−442. (2) Ai, H.; Jones, S. A.; Lvov, Y. M. Biomedical applications of electrostatic layer-by-layer nano-assembly of polymers, enzymes, and nanoparticles. Cell Biochem. Biophys. 2003, 39, 23−43. (3) Liang, Z.; Dzienis, K. L.; Xu, J.; Wang, Q. Covalent Layer-byLayer Assembly of Conjugated Polymers and CdSe Nanoparticles: Multilayer Structure and Photovoltaic Properties. Adv. Funct. Mater. 2006, 16, 542−548. (4) Kharlampieva, E.; Kozlovskaya, V.; Sukhishvili, S. A. Layer-byLayer Hydrogen-Bonded Polymer Films: From Fundamentals to Applications. Adv. Mater. 2009, 21, 3053−3065. (5) Sukhishvili, S. A.; Granick, S. Layered, erasable polymer multilayers formed by hydrogen-bonded sequential self-assembly. Macromolecules 2002, 35, 301−310.

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Chemistry of Materials (6) Such, G. K.; Johnston, A. P.; Caruso, F. Engineered hydrogenbonded polymer multilayers: from assembly to biomedical applications. Chem. Soc. Rev. 2011, 40, 19−29. (7) Johnston, A.; Read, E.; Caruso, F. DNA multilayer films on planar and colloidal supports: sequential assembly of like-charged polyelectrolytes. Nano Lett. 2005, 5, 953−956. (8) Johnston, A. P.; Caruso, F. Stabilization of DNA multilayer films through oligonucleotide crosslinking. Small 2008, 4, 612−618. (9) Kato, N.; Lee, L.; Chandrawati, R.; Johnston, A. P.; Caruso, F. Optically characterized DNA multilayered assemblies and phenomenological modeling of layer-by-layer hybridization. J. Phys. Chem. C 2009, 113, 21185−21195. (10) Inoue, H.; Sato, K.; Anzai, J. Disintegration of layer-by-layer assemblies composed of 2-iminobiotin-labeled poly (ethyleneimine) and avidin. Biomacromolecules 2005, 6, 27−29. (11) Cassier, T.; Lowack, K.; Decher, G. Layer-by-layer assembled protein/polymer hybrid films: nanoconstruction via specific recognition. Supramol. Sci. 1998, 5, 309−315. (12) Woolfson, D. N. The design of coiled-coil structures and assemblies. Adv. Protein Chem. 2005, 70, 79−112. (13) Yang, J.; Xu, C.; Wang, C.; Kopeček, J. Refolding hydrogels selfassembled from N-(2-hydroxypropyl) methacrylamide graft copolymers by antiparallel coiled-coil formation. Biomacromolecules 2006, 7, 1187−1195. (14) Aili, D.; Mager, M.; Roche, D.; Stevens, M. M. Hybrid Nanoparticle− Liposome Detection of Phospholipase Activity. Nano Lett. 2011, 11, 1401−1405. (15) Stevens, M. M.; Flynn, N. T.; Wang, C.; Tirrell, D. A.; Langer, R. Coiled-Coil Peptide-Based Assembly of Gold Nanoparticles. Adv. Mater. 2004, 16, 915−918. (16) Minelli, C.; Liew, J. X.; Muthu, M.; Andresen, H. Coiled coil peptide-functionalized surfaces for reversible molecular binding. Soft Matter 2013, 9, 5119−5124. (17) Stevens, M.; Allen, S.; Sakata, J.; Davies, M.; Roberts, C.; Tendler, S.; Tirrell, D.; Williams, P. pH-dependent behavior of surfaceimmobilized artificial leucine zipper proteins. Langmuir 2004, 20, 7747−7752. (18) White, S. J.; Morton, D. W. A.; Cheah, B. C.; Bronowska, A.; Davies, A. G.; Stockley, P. G.; Wälti, C.; Johnson, S. On-Surface Assembly of Coiled-Coil Heterodimers. Langmuir 2012, 28, 13877− 13882. (19) Aili, D.; Gryko, P.; Sepulveda, B.; Dick, J.; Kirby, N.; Heenan, R.; Baltzer, L.; Liedberg, B.; Ryan, M.; Stevens, M. Polypeptide folding-mediated tuning of the optical and structural properties of gold nanoparticle assemblies. Nano Lett. 2011, 11, 5564−5573. (20) Rydberg, J.; Baltzer, L.; Sarojini, V. Intrinsically unstructured proteins by designelectrostatic interactions can control binding, folding, and function of a helix-loop-helix heterodimer. J. Pept. Sci. 2013, 19, 461−469. (21) Aili, D.; Tai, F. I.; Enander, K.; Baltzer, L.; Liedberg, B. SelfAssembly of Fibers and Nanorings from Disulfide-Linked Helix− Loop−Helix Polypeptides. Angew. Chem., Int. Ed. 2008, 47, 5554− 5556. (22) Selegård, R.; Enander, K.; Aili, D. Generic phosphatase activity detection using zinc mediated aggregation modulation of polypeptidemodified gold nanoparticles. Nanoscale 2014, 6, 14204−14212. (23) Enander, K.; Aili, D.; Baltzer, L.; Lundström, I.; Liedberg, B. Alpha-helix-inducing dimerization of synthetic polypeptide scaffolds on gold. Langmuir 2005, 21, 2480−2487. (24) Stadler, B.; Chandrawati, R.; Goldie, K.; Caruso, F. Capsosomes: subcompartmentalizing polyelectrolyte capsules using liposomes. Langmuir 2009, 25, 6725−6732. (25) Chandrawati, R.; Städler, B.; Postma, A.; Connal, L. A.; Chong, S.-F.; Zelikin, A. N.; Caruso, F. Cholesterol-mediated anchoring of enzyme-loaded liposomes within disulfide-stabilized polymer carrier capsules. Biomaterials 2009, 30, 5988−5998. (26) Cui, J.; van Koeverden, M. P.; Müllner, M.; Kempe, K.; Caruso, F. Emerging methods for the fabrication of polymer capsules. Adv. Colloid Interface Sci. 2014, 207, 14−31.

(27) Chen, P.; Selegård, R.; Aili, D.; Liedberg, B. Peptide functionalized gold nanoparticles for colorimetric detection of matrilysin (MMP-7) activity. Nanoscale 2013, 5, 8973−8976. (28) Moutevelis, E.; Woolfson, D. N. A periodic table of coiled-coil protein structures. J. Mol. Biol. 2009, 385, 726−732.

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