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. 2024 Nov 2;15(1):9482.
doi: 10.1038/s41467-024-53555-0.

Atomically resolved imaging of the conformations and adsorption geometries of individual β-cyclodextrins with non-contact AFM

Márkó Grabarics #  1   2 , Benjamín Mallada #  3   4 , Shayan Edalatmanesh #  3   4 , Alejandro Jiménez-Martín  3   4   5 , Martin Pykal  4 , Martin Ondráček  3 , Petra Kührová  4 , Weston B Struwe  2   6 , Pavel Banáš  7 , Stephan Rauschenbach  8   9 , Pavel Jelínek  10   11 , Bruno de la Torre  12   13
Affiliations

Atomically resolved imaging of the conformations and adsorption geometries of individual β-cyclodextrins with non-contact AFM

Márkó Grabarics et al. Nat Commun. .

Abstract

Glycans, consisting of covalently linked sugar units, are a major class of biopolymers essential to all known living organisms. To better understand their biological functions and further applications in fields from biomedicine to materials science, detailed knowledge of their structure is essential. However, due to the extraordinary complexity and conformational flexibility of glycans, state-of-the-art glycan analysis methods often fail to provide structural information with atomic precision. Here, we combine electrospray deposition in ultra-high vacuum with non-contact atomic force microscopy and theoretical calculations to unravel the structure of β-cyclodextrin, a cyclic glucose oligomer, with atomic-scale detail. Our results, established on the single-molecule level, reveal the different adsorption geometries and conformations of β-cyclodextrin. The position of individual hydroxy groups and the location of the stabilizing intramolecular H-bonds are deduced from atomically resolved images, enabling the unambiguous assignment of the molecular structure and demonstrating the potential of the method for glycan analysis.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Fig. 1
Fig. 1. Structure and real-space images of β-cyclodextrin molecules by low-temperature STM and nc-AFM.
a Chemical formula of β-CD and side-view of a 3D-model highlighting the molecule’s truncated cone shape. b–d Nc-AFM images of β-CD on Au(111), recorded at constant height using frequency modulation detection. Representative examples are shown for the primary (b) and secondary face-up geometries (c), as well as for distorted molecules (d). e STM overview showing an irregular island formed upon electrospray deposition of β-CD onto Au(111) (bias = 1 V, tunneling current = 1 pA). The macrocycles stand out as doughnut-shaped structures from a lower background of fragments; the insert shows a higher resolution STM image of a single macrocycle (scale bar in the insert is 5 Å).
Fig. 2
Fig. 2. Comparison of experimental and simulated nc-AFM images of the β-cyclodextrin primary face.
a Atomic model of β-CD, representing the lowest-energy conformer at the DFT PBE level of theory, viewed from its primary face. More prominent functional groups are highlighted in color for ease of discussion. b Experimental and c simulated nc-AFM images of β-CD in the p↑ geometry, the tip-sample distance decreasing from left to right. In the experiment, z0 is defined as +250 pm relative to an initial STM setpoint above the molecular ring with the feedback at 1 V and 3 pA; Δz designates the difference in tip-sample distance relative to z0. d Overlay of atomic model and corresponding simulated image of β-CD in the p↑ geometry; the exact atomic groups giving rise to the various features can be readily identified. All scale bars are 5 Å.
Fig. 3
Fig. 3. Comparison of experimental and simulated nc-AFM images of the β-cyclodextrin secondary face.
a Atomic model of β-CD, representing the lowest-energy conformer at the DFT PBE level of theory, viewed from its secondary face. More prominent functional groups are highlighted in color. b Experimental and c simulated (bottom) nc-AFM images of β-CD in the s↑ geometry, the tip-sample distance decreasing from left to right. In the experiment, z0 is defined as +190 pm relative to an initial STM setpoint above the molecular ring with the feedback at 0.5 V and 1 pA; Δz designates the difference in tip-sample distance relative to z0. d Overlay of atomic model and corresponding simulated image of β-CD in the s↑ geometry. All scale bars are 5 Å.
Fig. 4
Fig. 4. Nc-AFM simulations and the electrostatic potential of β-cyclodextrin with its secondary face upwards.
The lateral relaxations of CO-like probe particles at tip-sample distances corresponding to the closest approach of the probe during the oscillation acquiring nc-AFM images shown in b and g are presented as red dots in panels a and f, respectively. Panels b–d hold the nc-AFM simulations without including the electrostatic potential, and sections g–i contain the simulations that include the electrostatic interaction between probe and sample. Panels e, j include the top (viewed from the secondary face) and side views of β-CD’s electrostatic potential map, respectively. The Δz values define the distance between the outermost atom of the molecule and the probe particle along the z-axis (perpendicular to the surface plane).
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