Structural and mechanistic insights into ABC transporters

Lead Research Organisation: University of Leeds
Department Name: Ctr for Plant Sciences

Abstract

Background: ABC transporters couple ATP hydrolysis to transmembrane movement of molecules. They are found in all organisms and transport substrates as diverse as ions and proteins. ABC transporters are grouped into families based on sequence homology. The members of the 'D' family occur principally in peroxisomes in fungi, plants and animals, although there is a plant protein in the chloroplast. In humans, mutations in ABCD1 lead to the severe X-linked disorder adrenoleukodystrophy, where very long chain fatty acids accumulate throughout the body, but particularly in myelin. In the model plant Arabidopsis Atabcd1 mutants fail to mobilise storage oil and have multiple biochemical and developmental abnormalities. In humans ABCD transporters are "half transporters" comprising a nucleotide binding domain and a set of 6 transmembrane domains and homodimerise to form the active transporter whereas in plants there is a single polypeptide that forms a fused heterodimer. They have a novel mechanism that involves cleavage of their acyl CoA substrates.
Objectives: 1) To explore the structure of AtABCD1 using a combination of EM and crystallography 2) To use Cys labelling combined with mass spectrometry and EM to explore conformational changes.
Novelty: Although there are high resolution structures for ABC transporters, there are none at all for any member of subfamily D despite their medical and agricultural importance.
Timeliness: Building on BBSRC funded research we have developed robust protocols for protein expression, purification and reconstitution at levels amenable for structural studies
Experimental approach: Protein expression and purification; crystallization; EM; protein labelling and mass spectrometry.

Publications

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Studentship Projects

Project Reference Relationship Related To Start End Student Name
BB/M011151/1 30/09/2015 29/09/2023
1941113 Studentship BB/M011151/1 30/09/2017 31/12/2021 Jack Wright