To each their own: Overcoming challenges in structural characterisation of closely related protein-protein complexes using single particle cryo-EM

Hagen Sülzen1,2, Sami Kereiche1, Jitka Votrubova1, Petr Pompach3, Sebastian Zoll1

1Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Flemingovo namesti 2, 16610 Prague 6, Czech Republic

2Charles University, Faculty of Science, Albertov 6, 12800 Prague 2, Czech Republic

3Charles University, Faculty of Science, Mass Spectroscopy, BIOCEV, Vestec, Czech Republic

With an increasing demand for structural characterisation of large protein-protein complexes, single-particle cryo-EM has been gaining popularity over the recent years. Due to widespread access to high powered microscopes and the continuous development of user-friendly data processing software, the main bottlenecks limiting the success of structure determination by cryo-EM, are now sample preparation and vitrification. Here we describe the technical challenges we faced during the structural characterisation of two closely-related protein-protein complexes, both entailing the same trypanosome surface protein but different naturally occurring variants of human complement factor 3.