Structural characterization of the NAP; the major adhesion complex of the human pathogen Mycoplasma genitalium

Abstract

Mycoplasma genitalium, the causative agent of non-gonococcal urethritis and pelvic inflammatory disease in humans, is a small eubacterium that lacks a peptidoglycan cell wall. On the surface of its plasma membrane is the major surface adhesion complex, known as NAP that is essential for adhesion and gliding motility of the organism. Here, we have performed cryo-electron tomography of intact cells and detergent permeabilized M. genitalium cell aggregates, providing sub-tomogram averages of free and cell-attached NAPs respectively, revealing a tetrameric complex with two-fold rotational (C2) symmetry. Each NAP has two pairs of globular lobes (named α and β lobes), arranged as a dimer of heterodimers with each lobe connected by a stalk to the cell membrane. The β lobes are larger than the α lobes by 20%. Classification of NAPs showed that the complex can tilt with respect to the cell membrane. A protein complex containing exclusively the proteins P140 and P110, was purified from M. genitalium and was structurally characterized by negative-stain single particle EM reconstruction. The close structural similarity found between intact NAPs and the isolated P140/P110 complexes, shows that dimers of P140/P110 heterodimers are the only components of the extracellular region of intact NAPs in M. genitalium.

Behaviors and Energy Source ofMycoplasma gallisepticumGliding

“…Previous studies show that one adhesin complex of M. pneumoniae is composed of two heterodimers, one of each is assembled by one P1 adhesin molecule and one P90 molecule (28). The adhesin complex in M. genitalium is also composed of a dimer of heterodimers constructed by P110 and P140, the homologs of P1 adhesin and P90, respectively (53). P110 has a binding site for SOs, so one adhesin complex binds two SOs (31).…”

Cryo‐electron tomography analyses of terminal organelle mutants suggest the motility mechanism of Mycoplasma genitalium

“…C, Supporting Information Fig. S2 and Movie S8), which showed the characteristic M. genitalium terminal organelle structure (Miyata and Hamaguchi, ) with the surface adhesion complexes covering the membrane protrusion (Scheffer et al ., ). This is consistent with our previous structural analysis depicting P140 and P110 as the only NAP extracellular components (Scheffer et al ., ).…”

“…The Mycoplasma proteins identified to play a role in the motility and adhesion have no homology to known bacterial or eukaryotic motility and adhesion proteins (Miyata, 2010). An example of a protein complex that is not part of the EDC but is essential for motility and adhesion is the surface adhesin (the so-called NAP) of M. genitalium, which we recently structurally characterized (Scheffer et al, 2017). However, the motility mechanism and the interplay between the NAPs and the EDC remain unclear (Seybert et al, 2006;Henderson and Jensen, 2006;Nakane et al, 2015).…”

Electron cryotomography of Mycoplasma pneumoniae mutants correlates terminal organelle architectural features and function

“…Kawamoto and colleagues () recently reported the markedly reduced abundance of these terminal organelle knobs on a mutant that lacks both the P1 adhesin and accessory proteins P40 and P90, concluding that these knobs correspond to the P1 adhesin complex. Moreover, isolated complexes of the P1/P90 homologues in the closely related Mycoplasma genitalium are structurally similar by ECT to the nap complexes seen on intact cells (Scheffer et al ., ). Our results here support and extend those findings, as tomograms of mutant III‐4, lacking P40 and P90 but not P1, likewise revealed visually lower numbers of protein knobs on the terminal organelle surface.…”

“…Thus P1, P40, and P90 are necessary but not sufficient for the presence of these terminal organelle knobs at wild‐type densities. Interestingly, Kawamoto and colleagues () noted a high degree of heterogeneity in the particles on wild‐type M. pneumoniae and suggested that they may be conformationally dynamic, while Scheffer and colleagues () described significant variation in the degree of tilt of nap complexes with respect to the membrane in M. genitalium .…”