Insights into the dynamics and actin interaction of the intrinsically disordered Tarp protein from Chlamydia

All Chlamydial species are prokaryotic, obligate, intracellular parasites which share a highly conservered host-cell internalisa-tion mechanism. The type-III secreted effector Tarp (Translocated actin recruiting protein) is essential during this process and mediates Chlamydial internalisation by nucleating host cell actin filaments to induce a phagocytosis-like encapsulation. Although Tarp is predicted to be intrinsically disordered, the minimal actin binding sequence from Tarp (100 residues) contains a short helix which shows homology with the well characterised WH2 actin binding helix. The presence of this potential helix localises Tarp along the Disordered:Ordered interface in charge/hydrophobic-ity phase space. Using nuclear magnetic resonance (NMR) spectroscopy and the minimal actin binding region, we have determined the residues affected by the Tarp:actin interaction. X-ray structures of other WH2:actin complexes show a short helix binding within the actin hydrophobic cleft and a region of 15-20 extended residues lying along the actin surface. Despite limited sequence homology it would seem that Tarp is binding actin with similar helical and extended elements. Based upon the affected residues of the 100 residue fragment, an 83 residue mutant has been synthesised which still contains the residues affected upon the Tarp:actin interaction. Furthermore, NMR spectra of both Tarp fragments alone indicates that in solution they exist as disordered polypeptides. T 1 and heteronuclear { 1 H-15 N} NOE relaxation studies of the fragments suggest that a region of decreased dynamics exists which corresponds to the WH2-like helix.