The Surface-Exposed Carboxyl Region of Mycoplasma pneumoniae Elongation Factor Tu Interacts with Fibronectin

Abstract

Mycoplasma pneumoniae is a bacterial pathogen of the human respiratory tract that causes a wide range of airway diseases as well as extrapulmonary symptoms. It possesses a distinct, differentiated terminal structure, termed the attachment organelle, that mediates adherence to the host respiratory epithelium. Previously, we reported that surface-associated M. pneumoniae elongation factor Tu (EF-Tu, also called MPN665) serves as a fibronectin (Fn)-binding protein, facilitating interactions between mycoplasmas and extracellular matrix. In the present study, we determined that binding of M. pneumoniae EF-Tu to Fn is primarily mediated by the EF-Tu carboxyl region. A 179-amino-acid region spanning the carboxyl terminus (designated EC; amino acids 192 to 394) binds Fn in a dose-dependent manner. Further analysis of carboxyl constructs (ED3 and ED4) and their deletion truncations (ED3.1, ED3.2, and ED4.1) revealed that the carboxyl region possessed two distinct sites with different Fn-binding efficiencies. Immunogold electron microscopy using antibodies raised against recombinant ED3 and ED4 demonstrated the surface accessibility of the EF-Tu carboxyl region. Competitive binding assays using intact radiolabeled mycoplasmas and purified recombinant ED3 and ED4 proteins, along with antibody blocking assays, reinforced the role of the surface-exposed EF-Tu carboxyl region in Fn binding. Alkali and high-salt treatment of mycoplasma membranes and Triton X-114-partitioned mycoplasma fractions confirmed the stable association of EF-Tu within the mycoplasma membrane. These observations highlight the unique, multifaceted, and unpredictable role of the classically defined cytoplasmic protein EF-Tu relative to cellular function, compartmentalization, and topography.

Selenium effects on the metabolism of a Se-metabolizingLactobacillus reuteri: analysis of envelope-enriched and extracellular proteomes

“…pneumoniae EF-Tu binds to fibronectin, 40 and in Lactobacillus johnsonii it is able to bind mucin and thus intestinal epithelial cells, also displaying immunomodulatory properties. 17 Another interesting up-regulated enzyme is bifunctional acetaldehyde-CoA/alcohol dehydrogenase (spot 4).…”

Anaplasma phagocytophilum Asp14 Is an Invasin That Interacts with Mammalian Host Cells via Its C Terminus To Facilitate Infection

“…Several recovered proteins are predicted to be cytoplasmic. While many of these socalled cytoplasmic proteins were recovered in another surface biotinylation and affinity proteomic study of A. phagocytophilum or have homologs in other bacteria that localize to and function on bacterial surfaces (47,(55)(56)(57)(58)(59)(60)(61)(62)(63)(64), it remains to be verified if all recovered proteins in this study are in fact surface localized. Though we initially identified Asp14 on the A. phagocytophilum DC surface, indirect immunofluorescence analyses revealed that the bacterium constitutively expresses Asp14 throughout its development in HL-60, RF/6A, and ISE6 cells.…”

“…APH_0240 (the chaperonin GroEL), APH_0346 (DnaK), and APH_1032 (the elongation factor Tu) were chosen because these housekeeping proteins have been shown to moonlight as surface proteins in A. phagocytophilum and other bacteria. Moreover, these proteins have been implicated as adhesins in other bacteria (47,(55)(56)(57)(58)(59)(60)(61)(62)(63)(64). Of the 11 candidates, Omp-1A, Omp85, Msp5, and APH_0441 carry predicted N-terminal signal peptide sequences.…”

Genomic and Surface Proteomic Analysis of the Canine Pathogen Staphylococcus pseudintermedius Reveals Proteins That Mediate Adherence to the Extracellular Matrix

“…The identification of intracellular proteins on the cell surface is not unexpected, and numerous studies of S. aureus have reported a similar phenomenon (10,16,46,53,58). In some cases, alternative functions associated with the cell surface have been described for cytoplasmic proteins, including GAPDH (26,52), enolase (4,23), transcription elongation factors (1,24), and chaperone proteins GroES and DnaK (20,57). In addition, ribosomal proteins are commonly identified on the bacterial cell surface and can exhibit a high degree of immunogenicity for humans (25,48).…”