Gliding motility of Mycoplasma mobile can occur by repeated binding to N-acetylneuraminyllactose (sialyllactose) fixed on solid surfaces

Abstract

Mycoplasma mobile relies on an unknown mechanism to glide across solid surfaces including glass, animal cells, and plastics. To identify the direct binding target, we examined the factors that affect the binding of Mycoplasma pneumoniae to solid surfaces and concluded that N-acetylneuraminyllactose (sialyllactose) attached to a protein can mediate glass binding on the basis of the following four lines of evidence: (i) glass binding was inhibited by N-acetylneuraminidase, (ii) glass binding was inhibited by N-acetylneuraminyllactose in a structure-dependent manner, (iii) binding occurred on glass pretreated with bovine serum albumin attached to N-acetylneuraminyllactose, and (iv) gliding speed depended on the density of N-acetylneuraminyllactose on glass.

FIG. 1.

Time course of cell binding in a tunnel chamber assay. Mycoplasma cells suspended in PBS-G with 10% horse serum were inserted into a tunnel chamber. The cells bound to the coverslip were counted at each time point, and the results are presented as a ratio relative to the total number of cells in the tunnel chamber.

FIG. 2.

Effects of serum on glass binding of cells of the wild type (WT) and the m13 mutant truncated for Gli349. Cells suspended in PBS-G containing various concentrations of serum were poured into a tunnel chamber, and the cells bound to the glass were counted after incubation for 600 s. The bound-cell ratio is presented relative to the total number of cells in the tunnel chamber.

FIG. 3.

Effects of N-acetylneuraminidase on glass binding and gliding speed. N-Acetylneuraminidase was added to cells gliding in a tunnel chamber, to 60 μg/ml. (A) Inhibition of glass binding. Numbers of bound cells are presented as ratios relative to the number at time zero. (B) Inhibition of gliding speed. Gliding speed is presented as a ratio relative to the initial gliding speed. Closed and open circles present results obtained with and without N-acetylneuraminidase, respectively.

FIG. 4.

Inhibitory effects of N-acetylneuraminyllactose on glass binding and gliding speed. (A) Number of cells bound to a glass surface after addition of 3′-N-acetylneuraminyllactose. The number of cells on the glass surface at each time point is presented as a ratio relative to that at time zero. (B) Gliding speed after addition of various concentrations of 3′-N-acetylneuraminyllactose. (C) Glass binding after addition of various concentrations of 6′-N-acetylneuraminyllactose. (D) Gliding speed after addition of various concentrations of 6′-N-acetylneuraminyllactose. The millimolar concentrations of N-acetylneuraminyllactose applied are indicated in panel D. The same set of concentrations was used for both compounds.

FIG. 5.

Binding of cells to glass pretreated with 3′-SLN-BSA or BSA. Coverslips were pretreated with various concentrations of 3′-SLN-BSA or BSA. Numbers of cells bound to the glass are presented as ratios relative to the total number of cells in the tunnel chamber. (A) Wild-type (WT) cells. (B) Strain m13 mutant cells.

FIG. 6.

Fluctuation of gliding speed at various densities of 3′-SLN-BSA. (A) Gliding speeds measured every 2 s on glass pretreated with various concentrations of 3′-SLN-BSA. (B) Gliding speeds measured every 0.1 s starting from 0.05 s earlier than the time indicated are presented by closed and open symbols for surfaces pretreated with 10 μg/ml 3′-SLN-BSA and 3 ng/ml 3′-SLN-BSA, respectively. Three traces from different cells are presented by different symbols for each condition. (C) Distribution of gliding speeds on glass pretreated with various concentrations of 3′-SLN-BSA. Averages and standard deviations are indicated by closed and open triangles, respectively.