Structural basis of receptor usage by the engineered capsid AAV-PHP.eB

Area of Science:

• Structural biology
• Gene therapy vectors
• Neuroscience

Background:

• Adeno-associated virus serotype 9 (AAV9) is a key vector for gene therapy, particularly for neurodegenerative diseases, due to its blood-brain barrier penetration.
• PHP.eB is a modified AAV9 variant engineered for enhanced central nervous system (CNS) potency.

Purpose of the Study:

• To determine the high-resolution cryo-electron microscopy structure of PHP.eB.
• To elucidate the structural basis for PHP.eB's enhanced CNS tropism and altered receptor interactions.

Main Methods:

• Cryo-electron microscopy (cryo-EM) at 2.24-Å resolution.
• Biochemical assays including pull-down experiments.
• Structural analysis of AAV variants and receptor interactions.

Main Results:

• The 7-amino acid insertion in PHP.eB adopts a distinct bent conformation, stabilized by lysine-aspartate interactions.
• Polycystic kidney disease 2 (PKD2) is identified as the primary AAV receptor (AAVR) domain for both AAV9 and PHP.eB.
• PHP.eB's structure shows a weakened interaction with AAVR and a potential additional interaction with LY6A, contributing to altered tropism.

Conclusions:

• The specific topology of the 7-mer insertion, influenced by internal residue interactions, dictates AAVR binding strength.
• Altered tropism of PHP.eB is attributed to both reduced AAVR binding and interaction with LY6A.
• Findings guide the design of future AAV libraries for optimized gene therapy vector selection and AAVR interaction modulation.