TY  - JOUR
A1  - Feiner, Rebecca Christine
A1  - Teschner, Julian
A1  - Teschner, Kathrin E.
A1  - Radukic, Marco T.
A1  - Baumann, Tobias
A1  - Hagen, Sven
A1  - Hannappel, Yvonne
A1  - Biere, Niklas
A1  - Anselmetti, Dario
A1  - Arndt, Katja Maren
A1  - Müller, Kristian Mark
T1  - rAAV Engineering for Capsid-Protein Enzyme Insertions and Mosaicism Reveals Resilience to Mutational, Structural and Thermal Perturbations
T2  - International journal of molecular sciences
N2  - 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 beta-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 beta-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.
KW  - adeno-associated-virus
KW  - beta-lactamase
KW  - inverted terminal repeat (ITR)
KW  - loop modification
KW  - capsid stability
Y1  - 2019
UR  - https://publishup.uni-potsdam.de/frontdoor/index/index/docId/47903
SN  - 1422-0067
VL  - 20
IS  - 22
PB  - MDPI
CY  - Basel
ER  -