Globotriaosylceramide (Gb(3)) is a glycosphingolipid present in the plasma membrane that is the natural receptor of the bacterial Shiga toxin. The unsaturation level of Gb(3) acyl chains has a drastic impact on lipid bilayer properties and phase behaviour, and on many Gb(3)-related cellular processes. For example: the Shiga toxin B subunit forms tubular invaginations in the presence of Gb(3) with an unsaturated acyl chain (U-Gb(3)), while in the presence of Gb(3) with a saturated acyl chain (S-Gb(3)) such invagination does not occur. We have used all-atom molecular dynamics simulations to investigate the effects of the Gb(3) concentration and its acyl chain saturation on the phase behaviour of a mixed bilayer of dioleoylphosphatidylcholine and Gb(3). The simulation results show that: (1) the Gb(3) acyl chains (longer tails) from one leaflet interdigitate into the opposing leaflet and lead to significant bilayer rigidification and immobilisation of the lipid tails. S-Gb(3) can form a highly ordered, relatively immobile phase which is resistant to bending while these changes for U-Gb(3) are not significant. (2) At low concentrations of Gb(3), U-Gb(3) and S-Gb(3) have a similar impact on the bilayer reminiscent of the effect of sphingomyelin lipids and (3) At higher Gb(3) concentrations, U-Gb(3) mixes better with dioleoylphosphatidylcholine than S-Gb(3). Our simulations also provide the first molecular level structural model of Gb(3) in membranes.