Mapping the Actin-Binding Region in the Tarp Protein from Chlamydia

Abstract

Chlamydia trachomatis is a gram negative obligate intracellular parasite responsible for the most common bacterial sexually transmitted disease in developed countries, and the leading cause of preventable blindness worldwide. Chlamydiae have a unique life cycle, with a metabolically inert, yet infectious, spore-like phase and a morphologically distinct metabolic phase inside the infected cell. During infection and multiplication, Chlamydiae affect the cellular processes in the host cell through a number of type-III secreted effectors. Early in the infection process, the 1005 residue protein Tarp (translocated actin recruiting protein) is injected into the host cell, where it causes remodelling of the host cell's actin cytoskeleton, forming an actin pedestal, which aids in the internalisation of the parasite. Tarp is predicted to be largely intrinsically disordered, and is capable of interacting directly with actin monomer to nucleate the formation of acting filaments. The minimal actin-binding region has been mapped to a 100-residue fragment, Tarp726-825 (Jewett et al (2006) PNAS, 103:15599-15604). We show that a shorter region of Tarp, Tarp726-808, is capable of binding actin and displays the characteristic NMR spectrum of a disordered protein. Using a combination of 2D and 3D nuclear magnetic resonance (NMR) experiments, the backbone resonances of this fragment were assigned. Comparison between {1H, 15N}-HSQC 2D NMR spectra in the presence and absence of actin reveals the amino acid residues directly affected by the binding.