[show abstract][hide abstract] ABSTRACT: MglA, a 22-kDa protein related to monomeric GTPases, is required for the normal operation of the A (Adventurous) and S (Social) motility and for multicellular development of Myxococcus xanthus. To determine how MglA controls A- and S-motility, MglA was assayed biochemically and its cellular location was determined. His-tagged MglA hydrolyzed GTP slowly in vitro at a rate nearly identical to that of Ras showing that MglA has GTPase activity. Immunofluorescence microscopy of fixed cells from liquid showed that MglA was associated with helical track similar to the MreB spiral that spanned the length of the cell. The distribution pattern of MglA depended on the type of surface from which cells were harvested. In cells gliding on 1.5% (w/v) agar, the helical pattern gave way to punctate clusters of MglA-Yfp at the poles and along the long axis (lateral clusters). The lateral clusters emerged near the leading pole as the cell advanced coincident with a decrease in the intensity of the MglA-Yfp cluster at the leading pole. Newly formed lateral clusters remained fixed with regard to the substratum as the cell moved forward, similar to focal adhesion complexes described for AglZ, a protein partner of MglA. Lateral clusters did not form in cells gliding in methylcellulose, a polymer that stimulates S-motility at low cell density; rather MglA-Yfp was diffuse in the cytoplasm and more concentrated at the poles. The results suggest that conditions that favor S-motility prevent the formation of lateral clusters of MglA, which are associated with A-motility functions.