SMOLBNET 2.0: Structural studies on proteins related to trypanosomatids infection and to complexes of these proteins with molecules which are involved in the infection process

Abstract

Many laboratories in the last 20 years, tried to identify proteins, on the surfase of Trypanosoma cruzi, which are involved in celular invasion. Conclusions from these works present a multistep process relating different molecules, both from the host and from the parasite, in a serie of events that produces movilization of Ca2+ in both cells. This project establish a colaboration between the Protein Crystallography group at IFSC/USP (EHR, HHMT e MD) and the Celular Biology and Parasite Inmunobiology laboratory at DMIP/UNIFESP (RM e DB), to use crystallographic structural determination and Structural Biology in the study of the trypanosomatides infection process. We defined two proteins for which, initial studies on their roll in the infection process have been performed, and the crystallographic structure should provide information on their biological function and on the consequences that inhibiting the protein should produce on the infective capacity of the parasites: A protein called P21 and mevalonate kinase (Mvak), both from Trypanosoma cruzi. The crystallographic studies that will produce the structure determination of these proteins have two main objectives: On one side, they will help to generate a detailed information on the biological function of these proteins. On the other side, in a longer time, they will be fundamental in generating a strategy of infection inhibition, though the design of specific inhibitors, that do not alter the activity of the homologous molecules of the host. Sequence homology studies allowed us to formulate the hypothesis that one of the domains of P21 have a folding corresponding to a type Kunitz Protease inhibitor. The crystallographic structure will clarify if this hypothesis is correct, and which are the aminoacids that have a high probability of being involved in interactions with the host. In the case of mevalonate kinase, from the detaild description of the structure, we will determine regions with structural differences with the human enzyme. The knowledge of these regions is the start point for the search of specific inhibitors for the trypanosome enzime. The membrane interacting proteins present in general a low solubility. Thus, it is necessary to adapt the crystallization search process, introducing detergents and a slight variation in temperature as tools to achieve the goal of obtaining high quality crystals to be used in structural determination. (AU)