High-speed atomic force microscopy directly visualizes conformational dynamics of the HIV Vif protein in complex with three host proteins

Vif (viral infectivity factor) is a protein that is essential for the replication of the HIV-1 virus. The key function of Vif is to disrupt the antiviral activity of host APOBEC3 (apolipoprotein B mRNA-editing enzyme catalytic subunit 3) proteins, which mutate viral nucleic acids. Inside the cell, Vif binds to the host cell proteins Elongin-C, Elongin-B, and core-binding factor subunit β, forming a four-protein complex called VCBC. The structure of VCBC–Cullin5 has recently been solved by X-ray crystallography, and, using molecular dynamics simulations, the dynamics of VCBC have been characterized. Here, we applied time-lapse high-speed atomic force microscopy to visualize the conformational changes of the VCBC complex. We determined the three most favorable conformations of this complex, which we identified as the triangle, dumbbell, and globular structures. Moreover, we characterized the dynamics of each of these structures. Our data revealed the very dynamic behavior of all of them, with the triangle and dumbbell structures being the most dynamic. These findings provide insight into the structure and dynamics of the VCBC complex and may support efforts to improve HIV treatment, because Vif is essential for virus survival in the cell.

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High-Speed AFM directly visualizes conformational dynamics of the HIV-Vif protein complex

10.1101/2020.05.18.102749 ◽

2020 ◽

Author(s):

Yangang Pan ◽

Luda S. Shlyakhtenko ◽

Yuri L. Lyubchenko

Keyword(s):

Conformational Changes ◽

Protein Complex ◽

High Speed ◽

Conformational Dynamics ◽

Md Simulations ◽

Time Lapse ◽

Hiv Treatment ◽

X Ray Crystallography ◽

Host Cell Proteins ◽

Virus Survival

AbstractViral infectivity factor (Vif) is a protein that is essential for the replication of the HIV-1 virus. The key function of Vif is to disrupt the antiviral activity of APOBEC3 proteins, which mutate viral nucleic acids. Inside the cell, Vif binds to the host cell proteins Elongin-C, Elongin-B, and CBF-β, forming a four-protein complex called VCBC. The structure of VCBC in complex with the Cullin5 (Cul5) protein has been solved by X-ray crystallography, and recently, using molecular dynamic (MD) simulations, the dynamics of VCBC and VCBC-Cul5 complexes were characterized. Here, we applied time-lapse high-speed atomic force microscopy (HS-AFM) to visualize the conformational changes of the VCBC complex. We determined the three most favorable conformations of the VCBC complex, which we identified as triangle, dumbbell, and globular structures. In addition, we characterized the dynamics of each of these structures. While our data show a very dynamic behavior for all these structures, we found the triangle and dumbbell structures to be the most dynamic. These findings provide insight into the structure and dynamics of the VCBC complex and support further research into the improvement of HIV treatment, as Vif is essential for virus survival in the cell.

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Faculty Opinions recommendation of Dynamics of nucleosomes assessed with time-lapse high-speed atomic force microscopy.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.13115957.14438056 ◽

2011 ◽

Author(s):

Yamini Dalal

Keyword(s):

Atomic Force Microscopy ◽

High Speed ◽

Time Lapse ◽

Force Microscopy ◽

Atomic Force

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Rigid-body fitting to atomic force microscopy images for inferring probe shape and biomolecular structure

PLoS Computational Biology ◽

10.1371/journal.pcbi.1009215 ◽

2021 ◽

Vol 17 (7) ◽

pp. e1009215

Author(s):

Toru Niina ◽

Yasuhiro Matsunaga ◽

Shoji Takada

Keyword(s):

Atomic Force Microscopy ◽

Rigid Body ◽

Correlation Coefficient ◽

High Speed ◽

Similarity Score ◽

Molecular Structures ◽

Cosine Similarity ◽

Force Microscopy ◽

Atomic Force ◽

Dynamics Simulations

Atomic force microscopy (AFM) can visualize functional biomolecules near the physiological condition, but the observed data are limited to the surface height of specimens. Since the AFM images highly depend on the probe tip shape, for successful inference of molecular structures from the measurement, the knowledge of the probe shape is required, but is often missing. Here, we developed a method of the rigid-body fitting to AFM images, which simultaneously finds the shape of the probe tip and the placement of the molecular structure via an exhaustive search. First, we examined four similarity scores via twin-experiments for four test proteins, finding that the cosine similarity score generally worked best, whereas the pixel-RMSD and the correlation coefficient were also useful. We then applied the method to two experimental high-speed-AFM images inferring the probe shape and the molecular placement. The results suggest that the appropriate similarity score can differ between target systems. For an actin filament image, the cosine similarity apparently worked best. For an image of the flagellar protein FlhAC, we found the correlation coefficient gave better results. This difference may partly be attributed to the flexibility in the target molecule, ignored in the rigid-body fitting. The inferred tip shape and placement results can be further refined by other methods, such as the flexible fitting molecular dynamics simulations. The developed software is publicly available.

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