Comprehensive AAV capsid fitness landscape reveals a viral gene and enables machine-guided design

Adeno-associated virus (AAV) capsids can deliver transformative gene therapies, but our understanding of AAV biology remains incomplete. We generated the complete first-order AAV2 capsid fitness landscape, characterizing all single-codon substitutions, insertions, and deletions across multiple funct...

Full description

Saved in:
Bibliographic Details
Published in:Science (American Association for the Advancement of Science) 2019-11, Vol.366 (6469), p.1139-1143
Main Authors: Ogden, Pierce J, Kelsic, Eric D, Sinai, Sam, Church, George M
Format: Article
Language:English
Subjects:
Biology
Capsid
Capsid Proteins - genetics
Capsids
Dependovirus - genetics
Empirical analysis
Fitness
Gene therapy
Genes, Viral
Genetic Therapy
Genomes
HEK293 Cells
Humans
Landscape design
Libraries
Membrane proteins
Mutagenesis
Mutagenesis, Insertional
Organs
Problem Solving
Protein engineering
Protein Engineering - methods
Proteins
Random mutagenesis
Saturation mutagenesis
Sequence Deletion
Thermal stability
Transfection
Viability
Viruses
VP1 protein
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Adeno-associated virus (AAV) capsids can deliver transformative gene therapies, but our understanding of AAV biology remains incomplete. We generated the complete first-order AAV2 capsid fitness landscape, characterizing all single-codon substitutions, insertions, and deletions across multiple functions relevant for in vivo delivery. We discovered a frameshifted gene in the VP1 region that expresses a membrane-associated accessory protein that limits AAV production through competitive exclusion. Mutant biodistribution revealed the importance of both surface-exposed and buried residues, with a few phenotypic profiles characterizing most variants. Finally, we algorithmically designed and experimentally verified a diverse in vivo targeted capsid library with viability far exceeding random mutagenesis approaches. These results demonstrate the power of systematic mutagenesis for deciphering complex genomes and the potential of empirical machine-guided protein engineering.
ISSN:0036-8075
1095-9203
DOI:10.1126/science.aaw2900