Unravelling the folding and stability of an ABC (ATP-binding cassette) transporter

PJ Booth, N Di bartolo

Research output: Contribution to journalArticle (Academic Journal)peer-review

6 Citations (Scopus)

Abstract

Prokaryotic importers from the large family of ABC (ATP-binding cassette) transporters comprise four separate subunits: two membrane-embedded and two cytoplasmic ATP-binding subunits. This modular construction makes them ideal candidates for studies of the intersubunit interactions of membrane protein complexes that contain both hydrophobic and hydrophilic subunits. In the present paper, we focus on the vitamin B12 importer of Escherichia coli, BtuCD, that contains two transmembrane BtuC subunits and two ATP-binding BtuD subunits. We have studied the factors that induce subunit dissociation and unfolding in vitro. The BtuCD complex remains intact in alcohol and mild detergents, but urea or SDS separate the BtuC and BtuD subunits, with 6 M urea causing 80% of BtuD to be removed from BtuCD. ATP is found to stabilize the complex as a result of its binding to the BtuD subunits. In the absence of ATP, low concentrations of urea (0.5–3 M) also induce some unfolding, with approximately 14% reduction in helicity in 3 M urea, whereas, in the presence of ATP, no changes are observed. Disassembly at the BtuD–BtuD dimeric interface in BtuCD can be achieved with smaller concentrations of urea (0.5–3 M) than that required to cause disassembly at the BtuC–BtuD transmission interface (3–8 M), suggesting a stronger interaction of the latter. The results also suggest that unfolding and disassociation of subunits appear to be coupled processes. Our work provides insights into the subunit interactions of an ABC transporter and lays the foundation for studies of the reassembly of BtuCD.
Translated title of the contributionUnravelling the folding and stability of an ABC (ATP-binding cassette) transporter
Original languageEnglish
Pages (from-to)751 - 760
Number of pages10
JournalBiochemical Society Transactions
Volume39 (3)
DOIs
Publication statusPublished - Jun 2011

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