Article Figures & Data
Figures
- FIG. 1.
Inhibitory effects of an antibody on gliding. Phase-contrast images of cells before (left) and 15 s after (right) addition of 300 μg/ml of the inhibitory antibody, obtained by superimposing video frames over a 2-s time span. The last frame was displayed in a different color. Bar, 5 μm.
- FIG. 2.
Immunoblot analysis of the wild-type (WT) strain and mutant m9 by inhibitory antibody. Whole-cell lysates were analyzed by SDS-5.5% polyacrylamide gel electrophoresis (left two lanes) staining with CBB and immunoblotting (right two lanes) using the anti-Gli521 monoclonal antibody (mAb-R19). The protein bands of Gli521 and Gli349 are indicated by open and hatched triangles, respectively. Molecular mass is indicated on the left.
- FIG. 3.
Gene arrangement around gliding genes in M. mobile and M. pulmonis. ORFs in the flanking regions of gli349 and gli521 were named from the predicted molecular weight. MYPU_2120, MYPU_2130, and MYPU_2140 form a single large ORF in M. pulmonis (ATCC 19612).
- FIG. 4.
Sequence of the gli521 gene. (A) Schematic of the gli521 gene. Predicted transmembrane segments (solid lines), nonsense mutation (asterisk), N terminus of mature protein (filled triangle), and epitope of antibody (hatched line) are shown. (B) Amino acid sequence near the N terminus. The predicted transmembrane segment, indicated by the box, is preceded by a cluster of lysine residues. The processed site is indicated by a triangle. (C) Nucleotide and amino acid sequences around the mutation point of m9. A codon, GAA, encoding glutamic acid, is changed to TAA, a stop codon.
- FIG. 5.
Subcellular localization of Gli521 and Gli349 proteins revealed by immunofluorescence microscopy. The cells were simultaneously stained for both proteins. (A) Fluorescence image detected for Gli521. (B) Merger of image shown in panel A with the phase-contrast image of panel F. (C) Fluorescence image detected for Gli349. (D) Merger of images shown in panels C and F. (E) Merger of images shown in panels A and C. (F) Phase-contrast image of cells. Bar, 2 μm.
- FIG. 6.
Subcellular localization of Gli521 by immunoelectron microscopy. Gli521 was labeled with 10-nm gold particles. The left two images are a stereo pair. Since some of the gold particles are difficult to see, they are marked by black dots in the right image at the positions at which they appear in the middle image. Most cells found on an electron microscopy grid showed a similar distribution of gold particles. A stereoscopic view was achieved by tilting the specimen 10 degrees. Bar, 0.2 μm.
- FIG. 7.
Inhibitory effects of anti-Gli521 antibody on gliding speed and glass binding. (A) Gliding speed after the addition of different amounts of anti-Gli521 antibody, shown as a function of time. The speed shown is the fraction of the initial speed, averaged over the cell population. (B) The number of cells bound to glass after the addition of different amounts of antibody, shown as a function of time. The number shown is the fraction of the initial number. The number of cells bound to a glass surface with an area of 3,200 μm2 was counted. The initial number of cells was more than 100. Cells were treated with 0 (⧫), 1 (□), 3 (▪), 10 (▵), 30 (▴), 100 (○), and 300 (•) μg/ml antibody.
- FIG. 8.
Inhibitory effects of anti-Gli521 antibody on the adsorption of cells to glass. The number of cells bound to a glass surface with an area of 3,200 μm2 was counted. (A) Inhibitory effects on glass binding of antibody added before or after glass binding of mycoplasma cells. Cells in suspension were mixed with various concentrations of antibody, inserted into a tunnel slide, and then the numbers of cells bound to glass were counted (filled circles), or cells bound to glass in a tunnel slide were exposed to various concentrations of antibody, and the numbers of cells bound to the glass were counted (open circles). The fraction of cells remaining on the glass is shown for each concentration. (B) Inhibitory effect of anti-Gli521 antibody on the subsequent removal of cells from glass by anti-Gli349 antibody. The fraction of cells remaining on the glass is shown for each time point.
- FIG. 9.
One possible mechanical cycle to explain the effects of antibody. A mycoplasma cell is represented by a hatched area bounded by a gray line. A leg mainly composed of a Gli349 molecule is represented as a rod sticking out from the cell, driven by or through Gli521 represented by an ellipse on the cell. The glass surface is represented by a solid line. The cyclic movement of the leg, including binding, stroke, and release, propels the cell body. The leg is shown differently for free, bound, and tightly bound forms. The binding of anti-Gli521 antibody inhibits step iii or iv and stops the leg, fixing the mycoplasma on the glass.
