Structure and tropism of wild-type AAV and of recombinant AAV vectors. (A) Gene therapy vectors are complex therapeutics requiring proper assembly of both DNA and protein components to generate the final product. Wild-type AAVs are small nonenveloped viruses, 20 to 25 nm in diameter, with a single-stranded DNA genome of ∼4.7 kb encoding 2 sets of genes, the rep genes required for replication and virion assembly and the cap genes that encode the 3 proteins that assemble to form the 60-mer viral capsid (upper bar). AAV vectors are composed of an outer protein shell, an exact or close replica of the AAV viral capsid, carrying a therapeutic gene of interest under the control of an appropriate promoter (lower bar). The vector is 74% protein by molecular weight. Maximum packaging capacity is ∼5 kb DNA, a limitation of AAV as a gene delivery vehicle. (B) Dozens of different naturally occurring AAV capsids, as well as genetically engineered ones, have been isolated for study, from humans and from other species. The capsid sequences are highly conserved, from 60% to >99%, but studies with naturally occurring serotypes⁵⁶  and purpose-engineered capsids^57-59  have shown that even small differences in capsid sequence may affect tissue tropism of a vector and can be exploited to improve therapeutic outcomes. Figure 1A reprinted from Xie et al⁹  with permission. Copyright 2000 National Academy of Sciences, USA. Figure 1B reprinted from Arrunda and Xiao⁶⁰  with permission. Copyright 2006 John Wiley and Sons.