University of Leeds, UK
Title: Kinetic and molecular dissection of coupled ion-substrate membrane transport proteins
Biography: Peter J F Henderson
The Mhp1 Na+-hydantoin membrane symport protein from Microbacterium liquefaciens is a paradigm for the nucleobase-cation-symport, NCS-1, family of transport proteins found widely in archaebacteria, bacteria, yeasts and plants. Their metabolic roles include the capture by cells of nitrogen compounds and vitamins from the environment. Mhp1 is also a structural model for the huge range of ‘5-helix-inverted-repeat’ superfamily of proteins, because, unusually, crystal structures are available for its open-outwards, occluded, and open-inward conformations. Here we accomplish a detailed dynamic model of the partial reactions in an alternating access cycle of membrane transport derived from substrate binding studies to the purified Mhp1 protein by combining novel mass spectrometry, stopped-flow and steady state kinetic analyses and mutagenesis. The mechanism of coupling substrate transport to the Na+-gradient is revealed during a sequence of mostly reversible kinetic steps that explain how transfer of substrate across the membrane is affected by changes in conformational states. The AceI H+/substrate antiport protein from Acinetobacter baumannii is a paradigm for the proteobacterial antimicrobial compound efflux (PACE) family of drug efflux proteins found dispersed throughout the Proteobacteria. AceI contributes to the resistance of Acinetobacter baumannii towards the widely used antiseptic, chlorhexidine. Currently there is little structural information about the PACE family of transport proteins, but progress towards understanding the recognition of substrates and cations by AceI and its homologues will be discussed.