Chemical introduction of reactive thiols into a viral nanoscaffold: a method that avoids virus aggregation.
The use of viral nanoparticles (VNPs) as building blocks for material fabrication has received particular attention in recent years. In earlier studies we showed the applicability of native gel electrophoresis in an agarose matrix as a useful method for the characterization of chemically modified VNPs. Here, we extend these studies and analyze the observed band pattern of intact Cowpea mosaic virus (CPMV) VNPs in agarose gels and show the applicability of native agarose gels for monitoring interparticle linkage of thiol-containing CPMV mutant particles. In addition, we report a protocol that allows the introduction of acetate-protected thiols to CPMV by means of a chemical reaction (rather than genetic modification). The advantage of this approach is that, by incorporating protected thiol groups, the formation of disulfide bonds leading to interparticle linkage is prevented. The resulting thiol-modified CPMV-SH(n) particles are stable, and following deprotection, the introduced thiols are reactive and can be labeled with thiol-selective reagents. They therefore provide a useful additional building block in the CPMV toolbox.
Steinmetz
Nicole F
NF
Department of Biological Chemistry, John Innes Centre, Colney, Norwich, Norfolk NR4 7UH, UK. nicole.steinmetz@bbsrc.ac.uk
Evans
David J
DJ
Lomonossoff
George P
GP
D017798:Comovirus / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry; D004587:Electrophoresis, Agar Gel / Q000379:methods; D005456:Fluorescent Dyes / Q000494:pharmacology; D008239:Lysine / Q000737:chemistry; D008956:Models, Chemical; D008958:Models, Molecular; D009154:Mutation; D053758:Nanoparticles / Q000737:chemistry*; D036103:Nanotechnology / Q000379:methods*; D012685:Sepharose / Q000737:chemistry; D012997:Solvents / Q000737:chemistry; D013438:Sulfhydryl Compounds / Q000737:chemistry*; D014461:Ultracentrifugation; D014780:Viruses / Q000378:metabolism*
D005456:Fluorescent Dyes; D012997:Solvents; D013438:Sulfhydryl Compounds; D012685:Sepharose; D008239:Lysine; D003545:Cysteine