Proteomic characterization of Legionella micdadei membranes

Legionella spp. are pathogens of free-living protozoa that can cause a respiratory disease called legionellosis. Pneumonia caused by L. micdadei is mainly associated with hospital infections and primarily affects immunosuppressed patients. The infection of host cells by L. micdadei is facilitated by specialized mechanisms through which the bacteria modulate host cell physiology to create a replicative niche. Central to this process is the Dot/Icm Type IV secretion system (T4SS), complemented by the Type II secretion system (T2SS) and other host-manipulation strategies involving specific proteins.

This study aimed to analyze the composition of the outer (OM) and inner membrane (IM) proteins isolated from L. micdadei bacteria.

The bacterial mass collected from BCYE plates was digested with DNase and RNase and then disintegrated in a French press. A sucrose density gradient was used to isolate the outer and inner membranes of L. micdadei. Proteins isolated from these membranes were analyzed using liquid chromatography with tandem mass spectrometry (LC-MS-MS). The L. micdadei proteome was investigated through diverse subtractive proteomics approaches, e.g., identification of pathogen-specific essential proteins, and draggability.

A detailed analysis of the L. micdadei membrane proteome allowed for the identification of 1784 proteins, including virulence factors, proteins associated with the transport of substances across cell membranes and the synthesis of cell membrane components, proteins related to bacterial mobility, and signal transduction pathways. The virulence factors present in L. micdadei membranes included protein building type IVB and II secretion systems, as well as major outer membrane protein (MOMP), macrophage infectivity potentiator (Mip), periplasmic protein EnhC, RND efflux system, outer membrane lipoprotein, the NodT family, and phosphatidylglycerol-prolipoprotein diacylglyceryl transferase.

The identification of L. micdadei membrane proteins and the elucidation of their functions provide deeper insights into the mechanisms underlying the pathogenesis of these microorganisms.