PEGylation of E2 virus-like nanoparticles

The structural core of the E2 subunit of B. stearothermophilus pyruvate dehydrogenase (E2) has been engineered to display antigens on its surface. For potential therapeutic applications, we have PEGylated a variant of E2 (E2-E279C) on introduced surface cysteines with methyl-PEG 24-maleimide. We verified PEGylation on the scaffold with mass spectrometry. Using dynamic light scattering and circular dichroism (CD), we showed the PEGylated proteins cage increased in size but the E2-E279C scaffold remained correctly folded. With CD thermostability test we confirmed that PEGylation dis not decrease the scaffold's high thermostability, with T m remaining near 90°C. Correct assembly was confirmed with transmission electron microscopy. We show that the designed E2-E279C mutant protein cage is chemically reactive toward thiol reactive compounds. Since application of E2-E279C protein cage enables easy and convenient "plug-and-play" surface modifications, we envision E2-E279C application not only as a drug delivery system, but also in imaging applications, cell targeting, scaffolds development for tissue engineering, and in vaccines design utilization.