Cryo-EM structures of the TolQ membrane protein in complex with inner membrane protein partner TolR from the opportunistic pathogen Acinetobacter baumannii provide information on structure and stoichiometry.

Abstract

TolQ-TolR-TolA is an energy-transducing inner-membrane sub-complex conserved across Gram-negative bacteria. It maintains connection with periplasmic TolB-Pal sub-complex in an energy-dependent manner through TolA protein spanning the periplasm. Utilizing proton motive force (PMF), Tol-Pal complex contributes to the integrity of outer membrane and the whole cell envelope. TolQ remains the only part of Tol-Pal complex with unknown structure up to now. To address this, we determined 3.02 and 3.34 Å cryo-electron microscopy (cryo-EM) structures of the integral inner-membrane protein TolQ and in complex with TolR, respectively from Acinetobacter baumannii. A. baumannii is one of the most dreaded and clinically important Gram-negative pathogens causing a broad-spectrum nosocomial infection mainly among debilitated patients admitted to intensive care units. Our structural analysis revealed that TolQ forms a pentamer with a channel-like hydrophobic trans-membrane pore slightly shifted above the membrane opening into a large negatively charged cytoplasmic chamber which is in common with the closest TolQ-TolR paralogs ExbB-ExbD and MotA-MotB. We validated observed structural features of TolQ by prior functional data. According to our analysis the most functionally detrimental point mutations mapped on structure are localized inside the channel. Obtained structures suggest that two trans-membrane domains of TolR dimer fit inside TolQ channel in TolQ-TolR complex forming a proton-flow pathway for PMF utilization. Cryo-EM structures solved in this work were funded in whole or in part with Refeyn Travel Grant and U.S. federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services, under contract No. HHSN272201700060C (Center for Structural Genomics of Infectious Diseases (CSGID); http://csgid.org).