title: The periplasmic membrane proximal domain of MacA acts as a switch in stimulation of ATP hydrolysis by MacB transporter.
abstract: Escherichia coli MacAB-TolC is a tripartite macrolide efflux transporter driven by hydrolysis of ATP. In this complex, MacA is the periplasmic membrane fusion protein that stimulates the activity of MacB transporter and establishes the link with the outer membrane channel TolC. The molecular mechanism by which MacA stimulates MacB remains unknown. Here, we report that the periplasmic membrane proximal domain of MacA plays a critical role in functional MacA-MacB interactions and stimulation of MacB ATPase activity. Binding of MacA to MacB stabilizes the ATP-bound conformation of MacB, whereas interactions with both MacB and TolC affect the conformation of MacA. A single G353A substitution in the C-terminus of MacA inactivates MacAB-TolC function by changing the conformation of the membrane proximal domain of MacA and disrupting the proper assembly of the MacA-MacB complex. We propose that MacA acts in transport by promoting MacB transition into the closed ATP-bound conformation and in this respect, is similar to the periplasmic solute-binding proteins.
authors.affiliation: Department of Chemistry and Biochemistry, University of Oklahoma, 101 Stephenson Parkway, Norman, OK 73019, USA.
authors.forename: Sita D
authors.initials: SD
authors.lastname: Modali
authors.forename: Helen I
authors.initials: HI
authors.lastname: Zgurskaya
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D019943:Amino Acid Substitution / Q000235:genetics; D000900:Anti-Bacterial Agents / Q000378:metabolism / Q000494:pharmacology; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D004917:Erythromycin / Q000378:metabolism / Q000494:pharmacology; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D006868:Hydrolysis; D026901:Membrane Transport Proteins / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D008954:Models, Biological; D020125:Mutation, Missense; D009827:Oleandomycin / Q000378:metabolism / Q000494:pharmacology; D011485:Protein Binding; D011487:Protein Conformation; D025941:Protein Interaction Mapping; D017434:Protein Structure, Tertiary
chemical_list: D018528:ATP-Binding Cassette Transporters; D000900:Anti-Bacterial Agents; D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; C436312:MacA protein, E coli; C436313:MacB protein, E coli; D026901:Membrane Transport Proteins; C073126:tolC protein, E coli; D004917:Erythromycin; D000255:Adenosine Triphosphate; D009827:Oleandomycin