STRUCTURAL AND FUNCTIONAL CHARACTERIZATION OF THE TRIPARTITE TRICARBOXYLATE TRANSPORT SYSTEM BctCBA OF Bordetella pertussis

Whooping cough, a respiratory disease caused by the Gram-negative bacterium Bordetella pertussis, poses a serious risk to unvaccinated infants and young children, potentially leading to respiratory complications and, in many cases, death. Although vaccination was introduced in 1940, the disease still accounts for approximately 24 million cases annually, and among children under five, it is estimated to cause approximately 160,000 deaths per year, in addition to persistent coughs of varying severity in older children, adolescents, and susceptible adults. Therefore, B. pertussis is considered a reemerging pathogen.

B. pertussis is a bacterium with strict nutritional requirements, as its primary carbon source is amino acids. Therefore, it utilizes highly efficient transport systems, notably the citrate transport system BctCBA, which belongs to the TTT (Tripartite Tricarboxylate Transporters) family. The BctCBA system forms a translocation channel composed of the transmembrane proteins BctB and BctA, while the BctC protein is the Solute Binding Protein (SBP). Interestingly, the genome of this bacterium overrepresents 78 genes corresponding to SBPs. Despite this, little is known about the role of this family of transporters in bacterial metabolism and their translocation mechanism.

Recent studies suggest that the BctCBA system transports citrate through an "elevator" mechanism, where the substrate is driven through the channel by cyclic movements. This mechanism allows the substrate-binding site to alternate between the cytoplasmic and periplasmic sides of the membrane, facilitating efficient citrate uptake even at low concentrations. The proposed project aims to elucidate the translocation mechanism in the BctCBA system using cryo-electron microscopy (SPA-cryo-EM) and crystallography. This will hopefully elucidate the function of the uptake system and contribute to potential targets for future therapeutic research focused on disease control.

This work is supported by São Paulo State Research Support Foundation (FAPESP) with processes 2022/11936-6 and 2024/20003-9