Controlling Supramolecular Complex Formation on the Surface of a

Christian Franceschini , Paolo Scrimin , and Leonard J. Prins ... Cristian Pezzato , Davide Zaramella , Massimiliano Martinelli , Grégory Pieters , M...
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Controlling Supramolecular Complex Formation on the Surface of a Monolayer-Protected Gold Nanoparticle in Water Grégory Pieters, Cristian Pezzato, and Leonard J. Prins* Department of Chemical Sciences, University of Padova, Via Marzolo 1, 35131 Padova, Italy S Supporting Information *

ABSTRACT: A combination of hydrophobic and electrostatic interactions drives the self-assembly of a large number of small molecules on the surface of a monolayer-protected gold nanoparticle. The hydrophobic interactions originate from the insertion of an aromatic unit in the hydrophobic part of the monolayer. This is evidenced by a shift in the emission wavelength of the fluorogenic probe upon binding. Up to around 35 small molecules can be simultaneously bound to the monolayer surface at micromolar concentrations in water. It is shown that an understanding of the supramolecular interactions that drive complex formation on the monolayer surface provides unprecedented control over the supramolecular chemistry occurring on the surface. By taking advantage of the different kinds of noncovalent interactions present in different probes, it is possibile to displace one type of surface-bound molecule from a heteromeric surface selectively. Finally, it is also possible to catch and release one type of surface-bound molecule selectively.

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INTRODUCTION The self-assembly of large supramolecular structures in water is an attractive approach for systems able to interact with biotargets such as proteins, oligonucleotides, and even cells.1 Self-assembly permits straightforward access to nanosized objects with minimal synthesis effort.2 Additional advantages include error correction during self-assembly and the adaptive behavior of the resulting structure with respect to the target or external stimuli (pH, temperature, metal ions, ...). Various supramolecular aggregates, such as micelles, rods, peptide amphiphiles, supramolecular polymers, and monolayer-protected nanoparticles, have been shown to be excellent structures for the recognition and sensing of biotargets.3−5 In particular, monolayer-protected gold clusters (Au MPCs) have attracted an enormous amount of attention because of their appealing intrinsic optical and electronic properties.6 This, in combination with a high biocompatibility, has led to numerous applications in nanomedicine7 and diagnostics.8 Inspired by the seminal contributions of Rotello et al.,9−11 we have recently started to develop an alternative approach to multivalent supramolecular systems relying on the self-assembly of small molecules on the surface of Au MPCs.12,13 The main advantage is that a heteromeric multivalent surface is simply generated by adding different small molecules to a single Au MPC scaffold. Compared to the formation of mixed monolayers on Au MPCs, this avoids the synthesis, purification, and characterization of each separate Au MPC, which is not always straightforward. Au MPCs are an attractive scaffold for this purpose because of their high stability even at nanomolar concentrations and their welldefined discrete dimensions. The use of noncovalent interactions for assembling the small molecules on the © 2012 American Chemical Society

monolayer surface in principle permits spontaneous adaptability to a target. A critical issue in this approach is the high affinity of the small molecules for the monolayer surface because complex formation should occur under saturation conditions even at low micromolar concentrations in water. Our studies have mainly involved Au MPC 1·Zn2+, which is a gold nanoparticle (d = 1.8 ± 0.4 nm) covered with hydrophobic C9-thiols terminated with a 1,4,7-triazacyclononane (TACN)·Zn2+ headgroup. This diameter implies that approximately 70 thiols are present on the surface of each nanoparticle.14 To ensure binding, small molecules were selected with the prerequisite of having a patch of negative charges in the form of either carboxylates or phosphates.15 Nonetheless, these studies provided clues that the binding affinity of the probes for the monolayer surface was not just caused by electrostatic interactions but also by interactions with the hydrophobic part of the monolayer.13 Similar observations have also been reported elsewhere.16−19 In general, many examples of supramolecular complex formation in water driven by a combination of hydrophobic and electrostatic interactions have been reported.1,20−22 Here, we provide compelling evidence that hydrophobic interactions play an important additional role in the binding of small molecules to Au MPC 1·Zn2+. The presence of a hydrophobic interaction permits a reduction of the number of negative charges in the probe without comprising the formation of the multivalent structure under saturation conditions. It is shown that the Special Issue: Interfacial Nanoarchitectonics Received: October 31, 2012 Revised: December 17, 2012 Published: December 21, 2012 7180

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Figure 1. Schematic representation of the formation of a multivalent supramolecular system and structures of the probes.

Figure 2. (a) Fluorescence intensities upon the addition of increasing amounts of dATPMANT to a solution of Au MPC 1·Zn2+ ([TACN·Zn2+] = 10 μM, [HEPES] = 10 mM, pH 7.0, T = 25 °C). On the right, the image shows the emission spectrum after the first additions. (b) Fluorescence intensities at 425 nm (red) and 448 nm (blue) as a function of the concentration of dATPMANT. (c) Maximum in the MANT emission wavelength (nm) as a function of the concentration of dATPMANT (red) superimposed on the fluorescence intensity at 448 nm (blue) as a function of the concentration of dATPMANT.

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possibility of regulating the binding strength of the probes gives an element of control over the supramolecular chemistry that takes place on the monolayer surface (selective exchange or catch and release of surface-bound small molecules).

RESULTS AND DISCUSSION

Electrostatic and Hydrophobic Interactions Drive Complex Formation on the Monolayer Surface. We decided to study the interaction of probe dATPMANT (2′-deoxy7181

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Figure 3. Surface saturation concentrations (μM) of ATPF and dATPMANT on Au MPC 1 in the presence and absence of Zn2+ at pH 7.0 and 8.0. The values for ATPF are taken from a study reported elsewhere.25

Figure 4. (a) Fluorescence intensities at 450 nm as a function of the concentration of dATPMANT, dADPMANT, or dAMPMANT added to Au MPC 1·Zn2+. Experimental conditions: [TACN·Zn2+] = 10 μM, [HEPES] = 10 mM, pH 7.0, T = 25 °C. (b) Normalized fluorescence intensities at 450 nm as a function of the concentration of ATP added to a solution of Au MPC 1·Zn2+ and either dATPMANT (dark blue), dADPMANT (intermediate blue), or dAMPMANT (light blue). Experimental conditions: [TACN·Zn2+] = 10 μM, [dATPMANT] = 3.0 μM, [dADPMANT] = [dAMPMANT] = 3.5 μM, [HEPES] = 10 mM, pH 7.0, T = 25 °C, slits 5/10. The solid lines are obtained from fitting the data to an appropriate model that was described before.12

observed (Figure 2b). Importantly, the observed maximum at 425 nm (23 nm below the expected wavelength) is a strong indication that the MANT fluorophore is inserted into the apolar monolayer. In fact, further additions of dATPMANT resulted in a strong increase in fluorescence emission with a maximum at 448 nm corresponding to the (expected) emission of free dATPMANT. As reported previously, from a plot of the fluorescence intensity at 448 nm as a function of the concentration of dATPMANT the surface saturation concentration (SSC) of the probe could be determined in a straightforward manner (SSCdATPmant = 3.2 μM). The SSC defines the number of molecules bound to the monolayer surface at saturation. The observation that the change in the maximum emission wavelength neatly occurs upon reaching the SSC confirms that it originates from a change in the environment of the MANT fluorophore (Figure 2c). It is interesting that just before reaching the SSC a small blue shift in the emission maximum is observed to 420 nm. This could point to an organizational process occurring on the surface to permit the accommodation of a major number of probes. The above experiments suggest that the MANT fluorophore is inserted into the monolayer. To obtain more information, we measured the interaction between dATPMANT and Au MPC 1·Zn2+ under conditions where no binding was observed for ATPF (2-aminopurine riboside-5′-O-triphosphate,28 λex = 305 nm, λem = 370 nm). ATPF is structurally very similar to dATPMANT but lacks the MANT fluorophore (Figure 1).

3′-O-(N′-methylanthraniloyl)adenosine-5′-O-triphosphate) with Au MPC 1·Zn2+ for several reasons. First, previous studies had already revealed a high affinity of ATP and its fluorescent analogue ATPF for Au MPC 1·Zn2+.15,23 Second, dATPMANT has an apolar aromatic unit attached to the deoxyribose that seemed perfectly placed to penetrate the monolayer. Third, the N-methylanthraniloyl (MANT) unit is fluorescent (λex = 355 nm, λem = 448 nm), which is essential for studying the binding to Au MPC 1·Zn2+. In addition, the MANT fluorophore is sensitive to its environment,24 and it is reported that the emission wavelength shifts 10−20 nm to shorter wavelengths in apolar environments. 25 Thus, the interaction between dATPMANT and Au MPC 1·Zn2+ was studied by means of a fluorescence titration taking advantage of the ability of gold nanoparticles to quench the fluorescence of bound fluorophores.26 All studies were performed in aqueous buffer ([HEPES] = 10 mM) at pH 7.0 with TACN headgroup concentrations equal to 10 ± 1 μM.27 A detailed analysis of the obtained spectra gave interesting insights into the nature of the complex formation. The first additions of dATPMANT resulted only in a minor fluorescence emission with a maximum at around 425 nm (Figure 2a). For these concentrations, the emission maximum and intensity were determined after subtracting the contribution from the signal at 407 nm owing to the Raman scattering of water (Supporting Information). Up to a concentration of around 2.5 μM dATPMANT, a linear increase in the (weak) emission intensity at this wavelength was 7182

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Figure 5. (a) Schematic representation of the selective exchange of WDDD from a heteromeric surface composed of WDDD and dATPMANT. (b) Normalized fluorescence intensities (based on that expected for the unbound probe) of WDDD and dATPMANT. Experimental conditions: [TACN·Zn2+] = 10 μM, [dATPMANT] = 1.0 μM, [WDDD] = 1.4 μM, [HEPES] = 10 mM, pH 7.0, T = 25 °C. (c) Schematic representation of the selective catch and release of WDDD from a heteromeric surface composed of WDDD and dATPMANT upon the removal and addition of Zn2+ from the monolayer. (d) Fluorescence intensities as a function of time upon the addition of Zn(NO3)2 (5 μM, downward arrow) or TPEN (5 μM, upward arrow) to a heteromeric surface composed of WDDD and dATPMANT. Experimental conditions: [TACN] = 10 μM, [dATPMANT] = 2.0 μM, [WDDD] = 1.0 μM;, [HEPES] = 10 mM, pH 7.0, T = 37 °C.

covering the gold nanoparticle,14 this implies that at saturation around 35 dAMPMANT probes are localized on the surface. Information on the relative affinity of the probes for the monolayer surface was obtained from a series of competition experiments between ATP and the MANT probes (Figure 4b). In these experiments, increasing amounts of the ATP competitor were added to a MANT probe bound to the surface of Au MPC 1·Zn2+, and the fluorescence emission from the released MANT probe was measured at 448 nm. These displacement experiments provide information on the relative affinity between ATP and the respective MANT probe. The displacement of 50% dAMPMANT (1.75 μM) required 4.0 μM of ATP corresponding to a relative binding affinity of around 2 in favor of dAMPMANT.31 The very similar binding affinity is remarkable considering the presence of just 2 negative charges in dAMPMANT against 4 for ATP. Another remarkable observation is that the addition of up to 1 mM ATP just replaced 0.75 μM dATPMANT (25%) from the surface. A control experiment in which ATP and dATPMANT were added in reverse order excluded the fact that these distributions were the result of kinetically slow exchange processes (Supporting Information). Controlling the Properties of a Dynamic Heteromeric Surface. The ability to regulate the binding affinity of small molecules to the monolayer surface is a key element in the application of these supramolecular structures as responsive systems. The peculiar high affinity of the MANT probes offers some important possibilities that will be illustrated by two examples. The first is the possibility to exchange one component from a heteromeric surface exclusively. A heteromeric surface was simply generated by adding dATPMANT (1.0 μM) and WDDD (1.4 μM) to Au MPC 1·Zn 2+ (Supporting Information). The carboxylate probe WDDD was chosen as a second unit because it has a much lower affinity compared to that of dATPMANT and because the fluorescence emission from tryptophan (λex = 280 nm, λem = 360 nm) can be monitored independently from the MANT fluorophore. The absence of fluorescence emission upon excitation of either

Previous studies had shown that in the absence of Zn2+ the SSC of ATPF decreased from 2.5 to 0.8 μM at pH 7.0, illustrating the importance of the metal ion in having a high SSC (Figure 3).29 Furthermore, at pH 8.0 a very low SSC value of 0.2 μM was measured for ATPF on Au MPC 1. On the contrary, dATPMANT showed nearly no decrease in SSC at pH 7.0 (2.8 μM) in the absence of Zn2+ and still a significant value of 1.8 μM at pH 8.0 (Figure 3). It is also noted that at pH 8.0 a shift in the emission wavelength was observed during the fluorescence titration (Supporting Information). So whereas for the ATPF probe the presence of Zn2+ (or, more generally, a positive charge) in the monolayer is essential to obtaining a significant SSC, these experiments show that this is much less relevant for dATPMANT. Although the SSC in itself does not provide information on the strength of the interaction (such information is obtained from the competition experiments described below), these results are indicative of a difference in the binding mode between ATPF and dATPMANT. For that reason, we extended our studies to dADPMANT and dAMPMANT. Interestingly, notwithstanding the reduced negative charge, both compounds still bound Au MPC 1·Zn2+ under saturation conditions at a headgroup concentration of 10 μM (Figure 4a). In particular for dAMPMANT, having just one phosphate group, this is impressive, especially considering the relatively low affinity of analogous compound AMP.15 Also for dADPMANT and dAMPMANT, a shift in the emission wavelength was observed during the fluorescence titrations, indicating that within the MANT series the difference in emission wavelength progressively decreased (Supporting Information, 23 nm for dATPMANT, 20 nm for dADPMANT, and 11 nm for dAMPMANT). This is tentatively ascribed to a different position of the MANT fluorophore in the hydrophobic part of the monolayer. It is important to note that a reduced size of the probe is accompanied by an increase in SSC (3.2 μM for dATPMANT, 4.2 μM for dADPMANT, and 4.8 μM for dAMPMANT). For dAMPMANT, this means a binding stoichiometry of 1 probe for each 2 TACN·Zn2+ complexes.30 With an estimated 70 thiols 7183

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Langmuir MANT or tryptophan indicates that both probes are quantitatively bound to the monolayer surface. The addition of ADP as a competitor results in the exclusive displacement of WDDD from the surface as witnessed by the full recovery of the expected fluorescence of WDDD and the complete absence of fluorescence emission from dATPMANT (Figure 5a). This is significant progress compared to our previous studies in which a selectivity factor of just 7 between two different probes was achieved.12 The second example exploits the high affinity of dATPMANT for Au MPC 1 even in the absence of Zn2+. By taking advantage of a protocol reported recently,29 this generates the possibility to catch and release the WDDD probe exclusively from the same heteromeric surface used before. Therefore, the addition of N,N,N′,N′-tetrakis(2pyridylmethyl) ethylenediamine (TPEN, a strong chelator for Zn2+) to Au MPC 1·Zn2+ covered with dATPMANT (2.0 μM) and WDDD (1.0 μM) resulted in the removal of Zn2+ from the monolayer surface (Figure 5b). In the absence of Zn2+, the WDDD probe loses its affinity for the surface and is released in solution with a consequent increase in fluorescence intensity. However, the SSC of dATPMANT is hardly affected by the removal of Zn2+ (see above), and no release of dATPMANT was observed at all. After the complete release of WDDD from the surface, Zn(NO3)2 was added, which remetalated the TACN headgroups, resulting in the capture of the WDDD probe. Two full catch-and-release cycles of WDDD were performed, and at no moment was any fluorescence emission of dATPMANT observed.

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REFERENCES

(1) Oshovsky, G. V.; Reinhoudt, D. N.; Verboom, W. Supramolecular chemistry in water. Angew. Chem., Int. Ed. 2007, 46, 2366−2393. (2) Lehn, J. M. Supramolecular chemistry - scope and perspectives, supermolecules, and molecular devices. Angew. Chem., Int. Ed. Engl. 1988, 27, 89−112. (3) Palmer, L. C.; Stupp, S. I. Molecular self-assembly into onedimensional nanostructures. Acc. Chem. Res. 2008, 41, 1674−1684. (4) Lim, Y. B.; Moon, K. S.; Lee, M. Recent advances in functional supramolecular nanostructures assembled from bioactive building blocks. Chem. Soc. Rev. 2009, 38, 925−934. (5) Uhlenheuer, D. A.; Petkau, K.; Brunsveld, L. Combining supramolecular chemistry with biology. Chem. Soc. Rev. 2010, 39, 2817−2826. (6) Daniel, M. C.; Astruc, D. Gold nanoparticles: assembly, supramolecular chemistry, quantum-size-related properties, and applications toward biology, catalysis, and nanotechnology. Chem. Rev. 2004, 104, 293−346. (7) Giljohann, D. A.; Seferos, D. S.; Daniel, W. L.; Massich, M. D.; Patel, P. C.; Mirkin, C. A. Gold nanoparticles for biology and medicine. Angew. Chem., Int. Ed. 2010, 49, 3280−3294. (8) Saha, K.; Agasti, S. S.; Kim, C.; Li, X. N.; Rotello, V. M. Gold nanoparticles in chemical and biological sensing. Chem. Rev. 2012, 112, 2739−2779. (9) De, M.; Rana, S.; Akpinar, H.; Miranda, O. R.; Arvizo, R. R.; Bunz, U. H. F.; Rotello, V. M. Sensing of proteins in human serum using conjugates of nanoparticles and green fluorescent protein. Nat. Chem. 2009, 1, 461−465. (10) Ghosh, P. S.; Verma, A.; Rotello, V. M. Binding and templation of nanoparticle receptors to peptide alpha-helices through surface recognition. Chem. Commun. 2007, 2796−2798. (11) You, C. C.; Miranda, O. R.; Gider, B.; Ghosh, P. S.; Kim, I. B.; Erdogan, B.; Krovi, S. A.; Bunz, U. H. F.; Rotello, V. M. Detection and identification of proteins using nanoparticle-fluorescent polymer 'chemical nose' sensors. Nat. Nanotechnol. 2007, 2, 318−323. (12) Pieters, G.; Cazzolaro, A.; Bonomi, R.; Prins, L. J. Self-assembly and selective exchange of oligoanions on the surface of monolayer protected Au nanoparticles in water. Chem. Commun. 2012, 48, 1916− 1918. (13) Zaramella, D.; Scrimin, P.; Prins, L. J. Self-assembly of a catalytic multivalent peptide-nanoparticle complex. J. Am. Chem. Soc. 2012, 134, 8396−8399. (14) Hostetler, M. J.; Wingate, J. E.; Zhong, C. J.; Harris, J. E.; Vachet, R. W.; Clark, M. R.; Londono, J. D.; Green, S. J.; Stokes, J. J.; Wignall, G. D.; Glish, G. L.; Porter, M. D.; Evans, N. D.; Murray, R. W. Alkanethiolate gold cluster molecules with core diameters from 1.5 to 5.2 nm: core and monolayer properties as a function of core size. Langmuir 1998, 14, 17−30. (15) Bonomi, R.; Cazzolaro, A.; Sansone, A.; Scrimin, P.; Prins, L. J. Detection of enzyme activity through catalytic signal amplification with functionalized gold nanoparticles. Angew. Chem., Int. Ed. 2011, 50, 2307−2312. (16) Ellis, A. V.; Vjayamohanan, K.; Goswaimi, R.; Chakrapani, N.; Ramanathan, L. S.; Ajayan, P. M.; Ramanath, G. Hydrophobic anchoring of monolayer-protected gold nanoclusters to carbon nanotubes. Nano Lett. 2003, 3, 279−282. (17) Bonomi, R.; Selvestrel, F.; Lombardo, V.; Sissi, C.; Polizzi, S.; Mancin, F.; Tonellato, U.; Scrimin, P. Phosphate diester and DNA hydrolysis by a multivalent, nanoparticle-based catalyst. J. Am. Chem. Soc. 2008, 130, 15744−15745. (18) Phillips, R. L.; Miranda, O. R.; Mortenson, D. E.; Subramani, C.; Rotello, V. M.; Bunz, U. H. F. Gold nanoparticle-PPE constructs as biomolecular material mimics: understanding the electrostatic and

CONCLUSIONS These data show that hydrophobic interactions play an important role in determining the affinity of small molecules for the surface of a cationic monolayer. The insertion of the MANT unit into the monolayer was evidenced by a blue shift in the emission wavelength. The presence of the hydrophobic unit permitted a reduction of the number of negative charges required to form complexes under saturation conditions. The use of smaller molecules causes an increase in the SSC and, thus, the valency of the obtained supramolecular nanostructure. This is of importance for future applications of these supramolecular systems in the field of (bio)recognition. Furthermore, the ability to diversify the binding interactions of small molecules with the monolayer service is important to the development of dynamic and responsive surfaces. Subsequent studies will be aimed at studying the importance of the nanoparticle (size) and monolayer structure (length of the alkyl chains) on complex formation on the monolayer surface. ASSOCIATED CONTENT

S Supporting Information *

Surface saturation concentrations, fluorescence titrations, displacement studies, and control experiments. This material is available free of charge via the Internet at http://pubs.acs.org.

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ACKNOWLEDGMENTS

Financial support from the ERC (StG-239898) and COST (CM0703 and CM0905) is acknowledged.

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Notes

The authors declare no competing financial interest. 7184

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hydrophobic interactions. Soft Matter 2009, 5, 607−612 . A correction was published in Soft Matter 20095, 5042. (19) Liu, X.; Hu, Y.; Stellacci, F. Mixed-ligand nanoparticles as supramolecular receptors. Small 2011, 7, 1961−1966. (20) Davis, J. T.; Spada, G. P. Supramolecular architectures generated by self-assembly of guanosine derivatives. Chem. Soc. Rev. 2007, 36, 296−313. (21) Kubik, S.; Reyheller, C.; Stuwe, S. Recognition of anions by synthetic receptors in aqueous solution. J. Incl. Phenom. Macrocycl. Chem. 2005, 52, 137−187. (22) Liu, L.; Guo, Q. X. The driving forces in the inclusion complexation of cyclodextrins. J. Incl. Phenom. Macrocycl. Chem. 2002, 42, 1−14. (23) Bonomi, R.; Cazzolaro, A.; Prins, L. J. Assessment of the morphology of mixed SAMs on Au nanoparticles using a fluorescent probe. Chem. Commun. 2011, 47, 445−447. (24) Hiratsuka, T. New ribose-modified fluorescent analogs of adenine and guanine nucleotides available as subtrates for various enzymes. Biochim. Biophys. Acta 1983, 742, 496−508. (25) Jameson, D. M.; Eccleston, J. F. In Fluorescence Spectroscopy; Brand, L., Johnson, M. L., Eds.; Academic Press: San Diego, 1997; Vol. 278, pp 363−390. (26) Dulkeith, E.; Morteani, A. C.; Niedereichholz, T.; Klar, T. A.; Feldmann, J.; Levi, S. A.; van Veggel, F.; Reinhoudt, D. N.; Moller, M.; Gittins, D. I. Fluorescence quenching of dye molecules near gold nanoparticles: radiative and nonradiative effects. Phys. Rev. Lett. 2002, 89, 203002. (27) The concentration of TACN headgroups is determined as reported previously15 from kinetic titrations using either Zn(NO3)2 or Cu(NO3)2. The accuracy of the obtained concentration is 10 ± 1 μM. (28) McClure, W. R.; Scheit, K.-H. Enzyme kinetic parameters of the fluorescent ATP analogue, 2-aminopurine triphosphate. FEBS Lett. 1973, 32, 267−269. (29) Pieters, G.; Pezzato, C.; Prins, L. J. Reversible control over the valency of a nanoparticle-based supramolecular system. J. Am. Chem. Soc. 2012, 134, 15289−15292. (30) The 1:2 stoichiometry was independently confirmed by catalytic inhibition experiments using a protocol described before15 (Supporting Information). (31) These values are approximated because the probe is present at a concentration slightly below the SSC to ensure the complete absence of initial fluorescence. Consequently, initial additions of ATP may result in surface binding without displacing the probe. This is also the reason that the initial points were excluded from the fitting of the dAMPMANT curve.

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