Molecular Stiffening by Macrocycle Clustering

Abstract

Allosteric stiffening of a portion of a protein surface is a strategy used in nature to regulate protein oligomerization and provide crucial functions for cells. However, a similar strategy to selectively control part of a compound dynamics remains elusive. Here we show that cucurbit[n]uril (CB[n]) macrocycles can bind almost all portions of a tetratopic guest molecule, stiffening the different parts of the guest to different extents. "Host-guest" interactions were found to be instrumental in selectively "freezing" guest molecular motions. The combination of ¹H-NMR (1D, 2D), DOSY, VT-NMR, isothermal titration calorimetry (ITC), mass spectrometry and molecular modelling enabled to highlight the crucial role of cucurbit[8]uril (CB[8]) binding in the selective hardening of relevant portions of the guest molecule. Beyond implications for bioinspired systems mimicking control of a system dynamic to create a new function, this approach has relevance for improving room temperature phosphorescence, and could also be used to allosterically control organocatalysis in water.

Keywords: Cucurbituril; Macrocycles; Rigidification; Supramolecular; Tetratopic.

Figure 1

Structures of guest molecules and macrocyclic hosts used in this work (V: viologen, P: phenylene, A: aromatic group).

Figure 2

a) Full range and b) expanded 700–1700 m/z range of the ESI mass spectrum recorded for the VP‐A‐PV/CB[8]₄ mixture after ion separation in the mobility cell of the instrument to enhance the dynamic range of detection and hence allow the weak signals of the targeted 1:4 complex (emphasized in b) to be best detected in spite of the highly abundant individual species (shown in a). Three ionic forms of the 1:4 complex were observed depending of the number of adducted Na⁺, as supported by their isotopic patterns (in black) matching theoretical ones (upside down, in gray) and accurate mass data provided for the m/z 1273.2 species in inset.

Figure 3

¹H NMR spectra (500 MHz, D₂O) of a) VP‐A‐PV, b) the VP‐A‐PV•CB[8]₄ complex (300 K), and c) the corresponding Diffusion‐Ordered SpectroscopY (DOSY) spectrum. Note the curved double arrows tentatively accounting for the qualitative molecular motion associated to the colored fragments.

Figure 4

Radial Distribution Functions (RDF) corresponding to distances in the VP‐A‐PV•CB[8]₄ complex (H₂O, 300 K) between hydrogen atoms of one CB[8] (equatorial H atoms) toward oxygen atoms of one CB[8] rim of the three other CB[8] (the rims considered are the closest to H atoms, those susceptible to engage in C─H•••O hydrogen bonds (abscissas correspond to distances (r) in Å, and ordinates correspond to the g(r) function, structures are from the MD trajectory and representative of the complex and green arrows qualitatively illustrate to magnitude of CB[8]–CB[8] interactions between the relevant cucurbiturils.

Figure 5

a) Ellipsoid representation of the VP‐A‐PV•CB[8]₄ complex (H₂O, 300 K) accounting for local positional variabilities around mean atom positions as deduced from molecular dynamics (Chimera, scale factor = 1, smoothing level = 3). Note the relatively large volumes around atoms of CB[8] positioned on viologen stations reflecting positional variations, large volumes also for the side chains included in the two corresponding CB[8], and small volumes for atoms close to the center of the guest with quasi‐minimal volumes for “h” protons indicating that these protons experience minimal displacement during the 100 ns trajectory at 300 K. b) Distances between the barycenter of the guest and that of each CB[8]. While CB[8]₃ and CB[8]₄ stay on the viologens at ∼13 Å from the center of VP‐A‐PV, CB[8]₁ and CB[8]₂ stay complexed at ∼8 Å from the center of the guest.

Figure 6

¹H NMR spectra (500 MHz, D₂O) of a) the VP‐A‐PV•CB[7]₂ complex (300 K) before addition of CB[8], b), c), d), e) forming a new VP‐A‐PV•CB[7]₂•CB8]₂ complex (300 K). Note the gradual disappearance of proton “h” signal and the upfield shift of signals of protons corresponding to butyl groups (acetone: reference). Excess CB[7] was used to ensure quasi‐quantitative binding of 2 CB[7] on the 2 viologens and did not impact CB[8] binding as the additional CB[7] could not interact with the guest side‐arms. (•: signals corresponding to a small amount of the VP‐A‐PV•CB[8]₄ complex as shown in (f), ✱ is for the proposed “host–host” interactions)).

Figure 7

Variable‐temperature ¹H NMR spectra (500 MHz, D₂O) of the VP‐A‐PV•CB[7]₂•CB[8]₂ complex with ¹H NMR spectra of solutions of VP‐A‐PV at 278 K (top) and of VP‐A‐PV•CB[7]₂ at 350 K (bottom) for comparisons.

Figure 8

a) Proposed CB[7]‐CB[8] exchange on viologen stations of the two complexes VP‐A‐PV•CB[7]₂•CB[8]₂ and VP‐A‐PV•CB[8]₄ and b) part of the 500 MHz ROESY spectrum of a solution of VP‐A‐PV with 3 equiv. CB[7] and 3 equiv. CB[8] in D₂O (mixing time 400 ms).