The University of Southampton
University of Southampton Institutional Repository

Effect of charge reversal mutations on the ligand- and membrane-binding properties of liver fatty acid-binding protein

Davies, J.K., Hagan, R.M. and Wilton, D.C. (2002) Effect of charge reversal mutations on the ligand- and membrane-binding properties of liver fatty acid-binding protein The Journal of Biological Chemistry, 277, (50), pp. 48395-48402. (doi:10.1074/jbc.M208141200).

Record type: Article

Abstract

Liver fatty acid-binding protein (FABP) is able to bind to anionic phospholipid vesicles under conditions of low ionic strength. This binding results in the release of ligand, the fluorescent fatty acid analogue 11-dansylaminoundecanoic acid (DAUDA), with loss of fluorescence intensity (Davies, J. K., Thumser, A. E. A., and Wilton, D. C. (1999) Biochemistry 38,16932-16940). Using a strategy of charge reversal mutagenesis, the potential role of specific cationic residues in promoting interfacial binding of FABP to anionic phospholipid vesicles has been investigated. Cationic residues chosen included those within the a-helical region (Lys-20, Lys-31, and Lys-33) and those that make a significant contribution to the positive surface potential of the protein (Lys-31, Lys-36, Lys-47, Lys-57, and Arg-126). Only three cationic residues make a significant contribution to interfacial binding, and these residues (Lys-31, Lys-36, and Lys-57) are all located within the ligand portal region, where the protein may be predicted to exhibit maximum disorder. The binding of tryptophan mutants, F3W, F18W, and C69W, to dioleoylphosphatidylglycerol vesicles, containing 5 mol% of the fluorescent phospholipid dansyldihexadecanoylphosphatidylethanolamine, was monitored by fluorescence resonance energy transfer (FRET). All three mutants showed enhanced dansyl fluorescence due to FRET on addition of phospholipid to protein; however, this fluorescence was considerably greater with the F3W mutant, consistent with the N-terminal region of the protein coming in close proximity to the phospholipid interface. These results were confirmed by succinimide quenching studies. Overall, the results indicate that the portal region of liver FABP and specifically Lys-31, Lys-36, and Lys-57 are involved in the interaction with the interface of anionic vesicles and that the N-terminal region of the protein undergoes a conformational change, resulting in DAUDA release.

PDF wilton2_rae2008.pdf - Other
Restricted to Registered users only
Download (288kB)

More information

Submitted date: 9 August 2002
Published date: 11 October 2002

Identifiers

Local EPrints ID: 35581
URI: http://eprints.soton.ac.uk/id/eprint/35581
ISSN: 0021-9258
PURE UUID: 36aa49d3-b020-4146-8f0b-e190a45cbcd4

Catalogue record

Date deposited: 19 May 2006
Last modified: 17 Jul 2017 15:47

Export record

Altmetrics

Contributors

Author: J.K. Davies
Author: R.M. Hagan
Author: D.C. Wilton

University divisions

Download statistics

Downloads from ePrints over the past year. Other digital versions may also be available to download e.g. from the publisher's website.

View more statistics

Atom RSS 1.0 RSS 2.0

Contact ePrints Soton: eprints@soton.ac.uk

ePrints Soton supports OAI 2.0 with a base URL of http://eprints.soton.ac.uk/cgi/oai2

This repository has been built using EPrints software, developed at the University of Southampton, but available to everyone to use.

We use cookies to ensure that we give you the best experience on our website. If you continue without changing your settings, we will assume that you are happy to receive cookies on the University of Southampton website.

×