Analysis of the MRP8-MRP14 protein-protein interaction by the two-hybrid system suggests a prominent role of the C-terminal domain of S100 proteins in dimer formation

Calcium-binding S100 proteins are thought to play a central role in calcium -mediated signal transduction pathways. They consist of two helix-loop-heli x, calcium-binding EF-hand domains. A characteristic feature is their tende ncy to form homo- and/or heterodimeric complexes. This report presents for the first time a functional "in vivo" approach to the analysis of S100 prot ein dimerization. Using the two-hybrid system we analyzed the dimerization of MRP8 (S100A8) and MRP14 (S100A9), two S100 proteins expressed in myeloid cells. It is reported that the MRP8-MRP14 heteromer is the clearly preferr ed complex in both man and mouse. The ability to homodimerize, however, app ears to be restricted to the murine MRPs. Interaction analysis of chimeric murine/human MRP14 proteins indicates, that the C-terminal EF-hand domain p lays a prominent role in MRP8 MRP14 interaction and determines the specific ity of dimerization, Site-directed mutagenesis of four evolutionary conserv ed hydrophobic amino acids, which have been recently supposed to be essenti al for S100 protein dimerization, suggests that at least one of these, name ly the most N-terminal located residue, is not critical for dimerization.