TY - JOUR AB - Recombinant adeno-associated viruses (rAAV) provide outstanding options for customization and superior capabilities for gene therapy. To access their full potential, facile genetic manipulation is pivotal, including capsid loop modifications. Therefore, we assessed capsid tolerance to modifications of the structural VP proteins in terms of stability and plasticity. Flexible glycine-serine linkers of increasing sizes were, at the genetic level, introduced into the 587 loop region of the VP proteins of serotype 2, the best studied AAV representative. Analyses of biological function and thermal stability with respect to genome release of viral particles revealed structural plasticity. In addition, insertion of the 29 kDa enzyme β-lactamase into the loop region was tested with a complete or a mosaic modification setting. For the mosaic approach, investigation of VP2 trans expression revealed that a Kozak sequence was required to prevent leaky scanning. Surprisingly, even the full capsid modification with β-lactamase allowed for the assembly of capsids with a concomitant increase in size. Enzyme activity assays revealed lactamase functionality for both rAAV variants, which demonstrates the structural robustness of this platform technology. DA - 2019 DO - 10.3390/ijms20225702 KW - adeno-associated-virus KW - β-lactamase KW - inverted terminal repeat (ITR) KW - loop modification KW - capsid stability LA - eng IS - 22 PY - 2019 SN - 1422-0067 T2 - International Journal of Molecular Sciences TI - rAAV Engineering for Capsid-Protein Enzyme Insertions and Mosaicism Reveals Resilience to Mutational, Structural and Thermal Perturbations UR - https://nbn-resolving.org/urn:nbn:de:0070-pub-29391289 Y2 - 2024-11-24T00:19:48 ER -