Determining the structure of protein complexes is crucial for a deeper understanding of their function in biological systems. Therefore, specifying the interacting interface of complexes that forms retroviral proteins with host cell machinery is essential for understanding the individual steps in the life cycle of retroviruses. Traditional techniques including X-ray crystallography, cryo-electron microscopy and nuclear magnetic resonance (NMR) that are commonly used to determine protein structure are often limited by sample quantity, complex size and, not least, laboratory instrumentation. Hydrogen deuterium exchange mass spectrometry (HDX-MS) is an elegant complementary method to study the protein interaction interface. For single proteins and protein complexes, HDX-MS can be used to obtain information on structure, protein-protein or protein-ligand interaction sites, allosteric effects and conformational changes induced by posttranslational modifications. HDX-MS has the added advantage of not being limited by the size of proteins or protein complexes, is highly sensitive and is able to detect coexisting protein conformations.

Calmodulin (CAM) is a small, ubiquitous, and evolutionarily highly conserved protein regulating basic cellular functions in all eukaryotes. As well known, calmodulin interacts with structural components of retroviruses in their life cycle, namely with N-terminal domain of Gag structural polyprotein termed matrix protein (MA), which is responsible for many key steps in interaction of the retroviral particle with the cytoplasmatic membrane within the infected cell. However, the mechanism of interaction still remains unclear. For deeper clarification the MA-CAM interaction interface, myrMAPPHis construct (myristoylated N-terminal part of Gag polyprotein with preserved cleavage site for viral protease) of the Mason-Pfizer monkey virus (M-PMV) was used. The structure of the MA-CAM complex is been extensively studied in our laboratory group by NMR, cross linking mass spectrometry and also by HDX-MS. The data obtained by HDX-MS indicate strongly influence of calmodulin to the dynamics of several parts of myrMAPPHis, especially by changing the availability of residues in the region of the second and third helix and the loop between them. Moreover, the residues in these regions are also significant for interaction of MA with phospholipids on the membrane. Furthermore, the data also show that calmodulin also affects the availability of residues in the viral protease cleavage site region of the fifth helix.

According to the results, the interaction of myrMAPPHis with calmodulin and subsequently with phospholipids on the membrane may be the right stimulus for the myristoyl switch leading to the reorganization of the myrMAPPHis structure and its cleavage from the polyprotein precursor Gag as part of the maturation process.