title: Predicting the loading of virus-like particles with fluorescent proteins.
abstract: The virus-like particle (VLP) of the Cowpea Chlorotic Mottle Virus (CCMV) has often been used to encapsulate foreign cargo. Here we show two different rational design approaches, covalent and noncovalent, for loading teal fluorescent proteins (TFP) into the VLP. The covalent loading approach allows us to gain control over capsid loading on a molecular level. The achieved loading control is used to accurately predict the loading of cargo into CCMV VLP. The effects of molecular confinement were compared for the differently loaded VLPs created with the covalent method. We see that the loading of more than 10 fluorescent proteins in the 18 nm internal cavity of the CCMV capsid gives rise to a maximum efficiency of homo-FRET between the loaded proteins, as measured by fluorescence anisotropy. This shows that already at low levels of VLP loading molecular crowding starts to play a role.
authors.affiliation: Laboratory for Biomolecular Nanotechnology and ‡Nanobiophysics (NBP), MESA+Institute for Nanotechnology, University of Twente , P.O. Box 217, 7500 AE Enschede, The Netherlands.
authors.forename: W Frederik
authors.initials: WF
authors.lastname: Rurup
authors.forename: Fabian
authors.initials: F
authors.lastname: Verbij
authors.forename: Melissa S T
authors.initials: MS
authors.lastname: Koay
authors.forename: Christian
authors.initials: C
authors.lastname: Blum
authors.forename: Vinod
authors.initials: V
authors.lastname: Subramaniam
authors.forename: Jeroen J L M
authors.initials: JJ
authors.lastname: Cornelissen
mesh_terms: D003001:Cloning, Molecular; D008164:Luminescent Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010316:Particle Size; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D013499:Surface Properties; D019183:Tombusviridae / Q000737:chemistry*; D058425:Vaccines, Virus-Like Particle / Q000737:chemistry*
chemical_list: D008164:Luminescent Proteins; D011994:Recombinant Proteins; D058425:Vaccines, Virus-Like Particle