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A Quantitative Dot Blot Assay for AAV Titration and Its Use for Functional Assessment of the Adeno-associated Virus Assembly-activating Proteins

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Completion of the AAV Structural Atlas: Serotype Capsid Structures Reveals Clade-Specific Features

Mario Mietzsch¹, Ariana Jose¹, Paul Chipman¹

¹Department of Biochemistry and Molecular Biology, Center for Structural Biology, McKnight Brain Institute, College of Medicine, University of Florida, Gainesville, FL 32610, USA.

|January 16, 2021

Summary

This study determined the Adeno-associated virus (AAV) capsid structures for AAV7, AAV11, AAV12, and AAV13, completing the structural atlas. These findings provide insights into AAV capsid assembly and potential for gene therapy vector engineering.

Area of Science:

Structural biology
Virology
Biochemistry

Background:

Adeno-associated viruses (AAVs) are crucial for gene therapy vectors.
Previous studies determined capsid structures for most AAV serotypes.
A complete structural understanding is vital for optimizing gene delivery.

Purpose of the Study:

To determine the cryo-electron microscopy structures of AAV7, AAV11, AAV12, and AAV13.
To complete the structural atlas of known AAV serotype capsids.
To analyze clade-specific structural features for vector engineering.

Main Methods:

Cryo-electron microscopy (cryo-EM) for high-resolution imaging.
Three-dimensional (3D) image reconstruction.
Structural analysis of capsid proteins and surface loops.

Main Results:

Determined high-resolution structures for AAV7, AAV11, AAV12, and AAV13 capsids.
AAV7 is the first reported clade D structure; AAV11/AAV12 are in an unassigned clade; AAV13 is a clade C hybrid.
Identified conserved AAV features and clade-specific loop conformations.

Conclusions:

The structural atlas of AAV serotypes is now complete.
Clade-specific loop conformations offer targets for vector engineering.
These structures will aid in developing improved gene delivery vectors with enhanced targeting and efficiency.

Keywords:

AAV capsid cryo-EM gene delivery genome packaging serotype

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