A small-molecule-dependent vaccine

Important results from the project

Overexpression of coronavirus replicase proteins from plasmids in the heterologous bacterial (E. coli) host & subsequent purification proved more difficult than indicated in source publications. We cloned, transcribed in vitro & purified a known RNA substrate for extension by all of our replicase proteins (purified with & without fusion tags) but no activity was seen. We thus targeted T7 RNA pol instead & 4 clones, each with a different Trp mutation, were made for producing future results.

Expected long term effects

Live attenuated vaccines (LAVs) would be technically superior to inactive vaccines if they were safer. The long-term plan is to overcome this hurdle by enabling control of LAV replication with an exogenously-added unnatural, orally-available small molecule. This principle should enable cheap and rapid vaccination of the whole world’s population against numerous diseases instead of currently prioritizing vaccination of just the developed world.

Approach and implementation

Goals were (i) making a small-molecule-dependent COVID replicase & (ii) patenting this. Overexpression of the replicase proteins in a heterologous bacterial system from 4 available plasmids proved more difficult than indicated in 2 publications. No extension activity of our purified replicase was seen using an RNA substrate. These unexpected results necessitated subcloning & also targeting T7 RNA pol. But the 9 month project duration was insufficient for completion of these new plans.