RBD projects - Results

Chronological list - Längst Lab

Elisabeth successfully expressed the RNA binding domain of Nucleocapsid (11-20; Elisabeth Silberhorn).

• two batches, sufficient protein (>5ml), sufficient concentrated (500 uM)
• protein is active even for several freeze/thaw cycles - checked by MST, nanoDSF and DLS
• MST: RBD binding affinity towards ssRNA2 (26nt) ca. 2 uM

244 Enamine compounds that were selected by PharmAI - docked against the RNA binding interface of Nucleocapsid RBD - were Echo plated (2bind - Clemens)

RBD - MST inhibitor screen - does not work

Conclusion: fluorescentRBD + Inhibitor is the best way to measure the binding affinities of the inhbitors - the inhibitor screen does not work by MST

RBD - nanoDSF screens

RBD (5uM) + Enamine compounds (100 uM) - (11-20; Gernot Längst)

about 15% hits; Tm reduced by ca. 0.5 to 1°C

worked well - the identified 53 compounds were further anlysed by MST - RBD - MST inhibitor screen

Testing of inhibitors and results

see: List of RNA-binding inhibitors found

next steps

MST hits - check for effect in vivo and for effect in RNA binding (EMSA RBD+RNA; MST)