Conditions to minimize soft single biomolecule deformation when imaging with atomic force microscopy

Godon, Christian; Teulon, Jean‐Marie; Odorico, Michaël; Basset, Christian; Meillan, Matthieu; Vellutini, Luc; Chen, Shu‐wen W.; Pellequer, Jean‐Luc

doi:10.1016/j.jsb.2016.12.011

Cited by 13 publications

(16 citation statements)

References 59 publications

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“…Operational aspects are unexpected consequences of particular choices of experimental setups and they have been observed numerous times [78,79,80] since their first description [81]. The best-known operational aspect in NP size determination is the physical state of NPs.…”

Section: Discussionmentioning

confidence: 99%

On the Operational Aspects of Measuring Nanoparticle Sizes

et al. 2018

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Nanoparticles are defined as elementary particles with a size between 1 and 100 nm for at least 50% (in number). They can be made from natural materials, or manufactured. Due to their small sizes, novel toxicological issues are raised and thus determining the accurate size of these nanoparticles is a major challenge. In this study, we performed an intercomparison experiment with the goal to measure sizes of several nanoparticles, in a first step, calibrated beads and monodispersed SiO2 Ludox®, and, in a second step, nanoparticles (NPs) of toxicological interest, such as Silver NM-300 K and PVP-coated Ag NPs, Titanium dioxide A12, P25(Degussa), and E171(A), using commonly available laboratory techniques such as transmission electron microscopy, scanning electron microscopy, small-angle X-ray scattering, dynamic light scattering, wet scanning transmission electron microscopy (and its dry state, STEM) and atomic force microscopy. With monomodal distributed NPs (polystyrene beads and SiO2 Ludox®), all tested techniques provide a global size value amplitude within 25% from each other, whereas on multimodal distributed NPs (Ag and TiO2) the inter-technique variation in size values reaches 300%. Our results highlight several pitfalls of NP size measurements such as operational aspects, which are unexpected consequences in the choice of experimental protocols. It reinforces the idea that averaging the NP size from different biophysical techniques (and experimental protocols) is more robust than focusing on repetitions of a single technique. Besides, when characterizing a heterogeneous NP in size, a size distribution is more informative than a simple average value. This work emphasizes the need for nanotoxicologists (and regulatory agencies) to test a large panel of different techniques before making a choice for the most appropriate technique(s)/protocol(s) to characterize a peculiar NP.

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Section: Discussionmentioning

confidence: 99%

On the Operational Aspects of Measuring Nanoparticle Sizes

et al. 2018

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“…Our study registers similar observations, but we propose a different orientation possibility for individual subunits within the complex. Differences between our interpretations might be due to the surface properties of the a) complex, b) the HOPG/mica surface, c) sample preparation, or d) imaging in air or buffer, which affects the overall behavior of the molecule under the microscope(32, 50). Association analyses by ITC reveals that the binding of individual subunits is strong (~10 −9 M; single set of binding mode) but differs by a factor of 10 from the latest report (~10 −10 M) in which one of the subunits was immobilized, unlike our solution based label free evaluation(1).…”

Section: Discussionmentioning

confidence: 78%

“…This can be explained by overall negative charge carried by FXIII-A 2 dimer surface, which relies on positive electrostatic patches on the FXIII-B subunit to adhere to the mica surface in a complexed state ( Figure 1d ). The differences in height might be attributed to adsorption effects on the structure of the protein(31, 32). Wrapping of FXIII-B subunits around FXIII-A 2 dimer occurred from one side, giving the molecule a bi-partite appearance, suggesting partial asymmetry in the complex.…”

Section: Resultsmentioning

confidence: 99%

The plasma Factor XIII heterotetrameric complex structure: unexpected unequal pairing within a symmetric complex

Singh

Nazabal²,

Kaniyappan

et al. 2019

Preprint

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Factor XIII (FXIII) is a predominant determinant of clot stability, strength, and composition. Plasma FXIII circulates as a pro-transglutaminase with 2 catalytic A subunits and 2 carrierprotective B subunits in a heterotetramer (FXIII-A 2 B 2 ). FXIII-A 2 and -B 2 subunits are synthesized separately and then assembled in plasma. Following proteolytic activation by thrombin and calcium-mediated dissociation of the B-subunits, activated FXIII (FXIIIa) covalently cross-links fibrin, promoting clot stability. The zymogen and active states of the FXIII-A subunits have been structurally characterized; however, the structure of FXIII-B subunits and the FXIII-A 2 B 2 complex have remained elusive. Using integrative hybrid approaches including atomic force microscopy, cross-linking mass spectrometry, and computational approaches, we have constructed the first all-atom model of the FXIII-A 2 B 2 complex. We also used molecular dynamic simulations in combination with isothermal titration calorimetry to characterize FXIII-A 2 B 2 assembly, activation, and dissociation. Our data reveal unequal pairing of individual subunit monomers in an otherwise symmetric complex, and suggest this unusual structure is critical for both assembly and activation of this complex. Our findings enhance understanding of mechanisms associating FXIII-A 2 B 2 mutations with disease and have important implications for the rational design of molecules to alter FXIII assembly and/or activity to reduce bleeding and thrombotic complications.

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“…Our study registers similar observations, but we propose a different orientation possibility for individual subunits within the complex. Differences between our interpretations might be due to the surface properties of the following: (a) complex; (b) the HOPG/mica surface; (c) sample preparation; or (d) imaging in air or buffer, which affects the overall behavior of the molecule under the microscope [32,50]. The association analyses by ITC reveal that the binding of individual subunits is strong (~10 −9 M, single set of binding mode) but differs by a factor of 10 from the latest report (~10 −10 M) in which one of the subunits was immobilized, unlike our solution-based label-free evaluation [1].…”

Section: Ih Approaches Reveal a Unique Fxiii Complex Structurementioning

confidence: 79%

“…This can be explained by an overall negative charge carried by FXIII-A 2 dimer surface, which relies on positive electrostatic patches on the FXIII-B subunit to adhere to the mica surface in a complexed state ( Figure 1). The differences in height might be attributed to adsorption effects on the structure of the protein [31,32]. Wrapping of FXIII-B subunits around FXIII-A 2 dimer occurred from one side, giving the molecule a bi-partite appearance, suggesting partial asymmetry in the complex.…”

Section: Afm Topographs Indicates Complex Formation Restricts the Conmentioning

confidence: 99%

The Plasma Factor XIII Heterotetrameric Complex Structure: Unexpected Unequal Pairing within a Symmetric Complex

Singh

Nazabal²,

Kaniyappan

et al. 2019

Biomolecules

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Factor XIII (FXIII) is a predominant determinant of clot stability, strength, and composition. Plasma FXIII circulates as a pro-transglutaminase with two catalytic A subunits and two carrierprotective B subunits in a heterotetramer (FXIII-A 2 B 2 ). FXIII-A 2 and -B 2 subunits are synthesized separately and then assembled in plasma. Following proteolytic activation by thrombin and calcium-mediated dissociation of the B subunits, activated FXIII (FXIIIa) covalently cross links fibrin, promoting clot stability. The zymogen and active states of the FXIII-A subunits have been structurally characterized; however, the structure of FXIII-B subunits and the FXIII-A 2 B 2 complex have remained elusive. Using integrative hybrid approaches including atomic force microscopy, cross-linking mass spectrometry, and computational approaches, we have constructed the first all-atom model of the FXIII-A 2 B 2 complex. We also used molecular dynamics simulations in combination with isothermal titration calorimetry to characterize FXIII-A 2 B 2 assembly, activation, and dissociation. Our data reveal unequal pairing of individual subunit monomers in an otherwise symmetric complex, and suggest this unusual structure is critical for both assembly and activation of this complex. Our findings enhance understanding of mechanisms associating FXIII-A 2 B 2 mutations with disease and have important implications for the rational design of molecules to alter FXIII assembly or activity to reduce bleeding and thrombotic complications.

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Conditions to minimize soft single biomolecule deformation when imaging with atomic force microscopy

Cited by 13 publications

References 59 publications

On the Operational Aspects of Measuring Nanoparticle Sizes

On the Operational Aspects of Measuring Nanoparticle Sizes

The plasma Factor XIII heterotetrameric complex structure: unexpected unequal pairing within a symmetric complex

The Plasma Factor XIII Heterotetrameric Complex Structure: Unexpected Unequal Pairing within a Symmetric Complex

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