Identification of a 123-Kilodalton Protein (Gli123) Involved in Machinery for Gliding Motility of Mycoplasma mobile

Abstract

ABSTRACT Mycoplasma mobile glides on a glass surface in the direction of its tapered end by an unknown mechanism. Two large proteins, Gli349 and Gli521, were recently reported to be involved in glass binding and force generation/transmission, respectively, in M. mobile gliding. These proteins are coded tandemly with two other open reading frames (ORFs) in the order p123-gli349-gli521-p42 on the genome. In the present study, reverse transcriptase PCR analysis suggested that these four ORFs are transcribed cistronically. To characterize the p123 gene coding a 123-kDa protein (Gli123) of 1,128 amino acids, we raised polyclonal antibody against the Gli123 protein. Immunoblotting for Gli123 revealed that Gli123 was missing in a mutant strain, m12, which was previously isolated and characterized by a deficiency in glass binding. Sequencing analysis showed a nonsense mutation at the 523rd amino acid of the protein in the m12 mutant. Immunofluorescence microscopy with the polyclonal antibody showed that Gli123 is localized at the head-like protrusion's base, the cell neck, which is specialized for gliding, as observed for Gli349 and Gli521. Localization of the gliding proteins, Gli349 and Gli521, was disturbed in the m12 mutant, suggesting that Gli123 is essential for the positioning of gliding proteins in the cell neck.