Use this URL to cite or link to this record in EThOS: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.542833
Title: Functional characterisation of bacterial tripartite ATP-independent periplasmic (TRAP) transporters
Author: Mulligan, Christopher
ISNI:       0000 0004 2709 0771
Awarding Body: University of York
Current Institution: University of York
Date of Award: 2008
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Abstract:
Tripartite ATP-independent periplasmic (TRAP) transporters are extracytoplasmic solute receptor (ESR)-dependent secondary transporters that are widespread in bacteria, but not eukaryotes. TRAP transporters are composed of an ESR component and 2 unequally sized integral membrane components. The large membrane component has 12 predicted transmembrane helices (TMHs) and is thought to form the translocation channel. The small membrane component is composed of 4 TMHs and is of unknown, but essential function. TRAP transporters combine an ESR, which is normally associated with ATP-binding cassette (ABC) transporters, with the utilisation of electrochemical gradients across the membrane, using an undefined mechanism. Analysis of the data contained within the relational database, TRAPdb (www.trapdb.org), has revealed a number of new insights into TRAP transporter function, including the observation that marine-dwelling organisms have a propensity for high numbers of TRAP transporters - possibly indicating a role for Na+ in the transport cycle. Structural and functional analysis of TRAP transporters has been limited due to the recalcitrant nature of integral membrane proteins. The work presented here details the first overexpression and purification of the integral membrane proteins of a TRAP transporter. Using Escherichia coli and Lactococcus lactis expression systems, the integral membrane proteins from the E. coli TRAP transporter, YiaMNO, and the Haemophilus influenzae TRAP transporter, SiaPQM, have been expressed and purified. The entire sialic acid-specific TRAP transporter, SiaPQM, has been functionally reconstituted into liposomes. The energetic requirements of transport by SiaPQM have been elucidated revealing that a Na+ gradient in combination with an applied membrane potential are required to achieve maximal transport rates. Under normal conditions, SiaPQM is a unidirectional transporter, unlike most other secondary transporters, however, the presence of excess unliganded ESR is able to induce efflux of substrate from the proteoliposomes.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.542833  DOI: Not available
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