The Na+- pumping NADH Dehydrogenase: Catalytic Mechanism, Evolution and Role in Pathogenesis

The ability of pathogenic bacteria to generate a sodium gradient is essential for their survival in the host internal environment and to establish a successful infection. The gradient of sodium is used to support a large variety of homeostatic and energetic processes, including pH regulation, ATP synthesis, cell motility, uptake of nutrients, toxin extrusion, as well as the efflux of drugs. A large number of bacterial pathogens contain a specialized respiratory enzyme, the Na⁺-pumping NADH dehydrogenase (Na⁺-NQR), which is both the entry site of electrons into the respiratory chain and the main ion pump. The importance of Na⁺-NQR in the cell metabolism lies in the fact that its activity links the operation of the entire metabolic network with the mechanisms used to sustain ion homeostasis. Indeed, previous reports have demonstrated that Na⁺-NQR inactivation decreases the infectivity of bacterial cells and its activity regulates the production of virulence factors. My current data now demonstrates that this enzyme is the major step controlling the respiration of Pseudomonas aeruginosa and Chlamydia trachomatis. Moreover, Na⁺-NQR inhibition abolishes completely the ability of C. trachomatis to infect HeLa cells.

In addition to its key metabolic role, the Na⁺-NQR complex presents several highly unusual biochemical characteristics, not present in other respiratory enzymes or ion transporters, which indicate that Na⁺-NQR has evolved in a unique pattern, separated from other protein families. Recent studies by our group have clarified several important aspects of Na⁺-NQR, such as the electron transfer pathways and the mechanisms used for sodium pumping, which has led us to propose a model that describes the operation of the enzyme. The data indicate that the transport of sodium depends on a novel mechanism that couples the electron transfer between the redox centers of the enzyme, with conformational changes that control the accessibility of sodium to the binding sites.

The phylogenetic analysis of four of the Na⁺-NQR subunits has provided a background to propose a comprehensive scenario to understand the main molecular events involved in the evolution and spreading of this enzyme. The data indicates that this respiratory complex appeared in the Chlorobi/ Bacteroidetes ancestor, through the duplication of a homologous operon, encoding a membrane -bound Na⁺- pumping ferredoxin: NADH dehydrogenase. Subsequently, the ancestral protein complex evolved into the modern Na⁺-NQR, through a series of gene loses, re-functionalizations of certain subunits and acquisition of another subunit, from the aromatic monooxygenase family. Due to several advantageous traits, the Na⁺-NQR complex was spread to other bacterial lineages, through multiple events of horizontal gene transfer events.