8a9u

Publication Abstract from PubMed

AAV virion biology is still lacking with respect to complete understanding of the role the various structural subunits (VP1, 2, and 3) play in virus assembly, infectivity, and therapeutic delivery for clinical indications. In this study, we focus on the less studied AAV3B and generate a collection of AAV plasmid substrates that assemble virion particles deficient specifically in VP1, VP2 or VP1 and 2 structural subunits. Using a collection of biological and structural assays, we observed that virions devoid of VP1, VP2 or VP1 and 2 efficiently assembled virion particles, indistinguishable by cryo-EM to that of wild type, but unique in virion transduction (wild type > VP2 > VP1 > VP1 and 2 mutants). We also observed that the missing structural subunit was mostly compensated by additional VP3 protomers in the formed virion particle. Using cryo-electron microscopy (cryo-EM) analysis, virions fell into three classes, namely full, empty and partially-filled based on comparison of density values within the capsid. Further, we characterise virions described as "broken" or "disassembled" particles and provide structural information that supports the particle dissolution occurring via the two-fold symmetry. Finally, we highlight the unique value of employing cryo-EM as an essential tool for release criteria with respect to AAV manufacturing.

Cellular and structural characterisation of VP1 and VP2 knockout mutants of AAV3B serotype and implications for AAV manufacturing.,Arriaga I, Navarro A, Etxabe A, Trigueros C, Samulski RJ, Moullier P, Francois A, Abrescia NGG Hum Gene Ther. 2022 Sep 9. doi: 10.1089/hum.2022.119. PMID:36082996^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.