Nucleotide sequence of the Escherichia coli motB gene and site-limited incorporation of its product into the cytoplasmic membrane

Abstract

The motB gene product of Escherichia coli is an integral membrane protein required for rotation of the flagellar motor. We have determined the nucleotide sequence of the motB region and find that it contains an open reading frame of 924 nucleotides which we ascribe to the motB gene. The predicted amino acid sequence of the gene product is 308 residues long and indicates an amphipathic protein with one major hydrophobic region, about 22 residues long, near the N terminus. There is no consensus signal sequence. We postulate that the protein has a short N-terminal region in the cytoplasm, an anchoring region in the membrane consisting of two spanning segments, and a large cytoplasmic C-terminal domain. By placing motB under control of the tryptophan operon promoter of Serratia marcescens, we have succeeded in overproducing the MotB protein.Under these conditions, the majority of MotB was found in the cytoplasm, indicating that the membrane has a limited capacity to incorpor.1te the protein. We conclude that insertion of MotB into the membrane requires the presence of other more hydrophobic components, possibly including the MotA protein or other components of the flagellar motor. The results further reinforce the concept that the total flagellar motor consists of more than just the basal body. At least five proteins are essential for motor rotation (41, 47), but none of these have been found within the flagellar basal body (1,. 20), even though the basal body is considered a major part of the motor. Two of these proteins, MotA and MotB, are integral to the cell membrane (6, 37, 41). They are not necessary for assembly of the flagellum and do not copurify with it (20). They can be synthesized after flagellar assembly and used to activate the motor; paralyzed motA motB mutants acquire the ability to rotate their flagella after MotA and MotB synthesis under the direction of lambda-E. coli hybrid bacteriophage (41). Recently, it was shown (6) that, at least in the case of MotB, this acquisition of motility proceeds by quantum increases in flagellar rotation rate, presumably as a result of successive incorporation of subunits of MotB protein. The fact that motA and motB null mutants are nonetheless flagellated makes these genes different from all other flagellum-associated genes and will have * Corresponding author.

Functional Reconstitution of the Na+-driven Polar Flagellar Motor Component of Vibrio alginolyticus

“…Two types of motors, proton-driven (2) and sodium-driven (4), dependent on different coupling ions, have been described. The proton-driven motors of Escherichia coli and Salmonella typhimurium have been extensively studied, and the stator part of the torque generator consists of two cytoplasmic membrane proteins, MotA and MotB, which contain four transmembrane domains and one transmembrane domain, respectively (5)(6)(7)(8). Much genetic and physiological evidence suggests that MotA and MotB together form a proton channel (9 -13), and this complex is believed to be anchored to the cell wall via the peptidoglycan-binding domain of MotB (8,14,15).…”

Gate-controlled proton diffusion and protonation-induced ratchet motion in the stator of the bacterial flagellar motor

“…Escherichia coli has a few proton-driven flagellar motors with stators (protein MotA/B complex) in the inner membrane that act as proton channels (1)(2)(3)(4)(5). In addition, Vibrio alginolyticus has a polar flagellum powered by sodium ions (6).…”

“…MotB (308 residues) is composed of a short N-terminal cytoplasimic segment, one TM helix (B), and a large C-terminal periplasmic domain (Fig. 1B) (4,5). Asp32, which is situated near the cytoplasmic end of the B segment, is conserved across the species and considered to be the most plausible proton binding site (25).…”

Complete genome sequence analysis of the fish pathogen Flavobacterium columnare provides insights into antibiotic resistance and pathogenicity related genes

“…Cells with mutations in any of gliding genes are completely nonmotile, and they form nonspreading colonies exhibit no movement on agar or glass surfaces. Another two MotBrelated proteins (Pf1_00564, Pf1_01212) were also found, known as the component of the flagellar motor driving the flagella rotation [52]. But, no genes encoding flagella, pili or any motility organelles were found in the F. columnare genome, indicating the two proteins may assist to swing or did other functions.…”