@article {1139,
	title = {Post-translational modifications in capsid proteins of recombinant adeno-associated virus (AAV) 1-rh10 serotypes [Mass Spectrometry Facility - Proteomics \& Glycomics].},
	journal = {FEBS J},
	year = {2019},
	month = {2019 Jul 22},
	abstract = {<p>Post-translational modifications in viral capsids are known to fine-tune and regulate several aspects of the infective life cycle of several viruses in the host. Recombinant viruses that are generated in a specific producer cell line are likely to inherit unique post-translational modifications during intra-cellular maturation of its capsid proteins. Data on such post-translational modifications in the capsid of recombinant adeno-associated virus serotypes (AAV1-rh10) is limited. We have employed liquid chromatography and mass spectrometry analysis to characterize post-translational modifications in AAV1-rh10 capsid protein. Our analysis revealed a total of 52 post-translational modifications in AAV2-AAVrh10 capsids, including ubiquitination (17\%), glycosylation (36\%), phosphorylation (21\%), SUMOylation (13\%) and acetylation (11\%). While AAV1 had no detectable post-translational modification, at least four AAV serotypes had \&gt;7 post-translational modifications in their capsid protein. About 82\% of these post-translational modifications are novel. A limited validation of AAV2 capsids by MALDI-TOF and western blot analysis demonstrated minimal glycosylation and ubiquitination of AAV2 capsids. To further validate this, we disrupted a glycosylation site identified in AAV2 capsid (AAV2-N253Q), which severely compromised its packaging efficiency (~100-fold vs AAV2 wildtype vectors). In order to confirm other post-translational modifications detected such as SUMOylation, mutagenesis of a SUMOylation site(K258Q) in AAV2 was performed. This mutant vector demonstrated reduced levels of SUMO-1/2/3 proteins and negligible transduction, two weeks after ocular gene transfer. Our study underscores the heterogeneity of post-translational modifications in AAV vectors. The data presented here, should facilitate further studies to understand the biological relevance of post-translational modifications in AAV life cycle and the development of novel bioengineered AAV vectors for gene therapy applications. This article is protected by copyright. All rights reserved.</p>
},
	issn = {1742-4658},
	doi = {10.1111/febs.15013},
	author = {Mary, Bertin and Maurya, Shubham and Arumugam, Sathyathithan and Kumar, Vikas and Jayandharan, Giridhara R}
}
