Tryptophan probes reveal residue-specific phospholipid interactions of apolipoprotein C-III.

Autor:	Pfefferkorn CM; Laboratory of Molecular Biophysics, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA., Walker RL 3rd; Laboratory of Molecular Biophysics, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA., He Y; Laboratory of Biochemistry, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA., Gruschus JM; Laboratory of Molecular Biophysics, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA., Lee JC; Laboratory of Molecular Biophysics, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA. Electronic address: leej4@mail.nih.gov.
Jazyk:	angličtina
Zdroj:	Biochimica et biophysica acta [Biochim Biophys Acta] 2015 Nov; Vol. 1848 (11 Pt A), pp. 2821-8. Date of Electronic Publication: 2015 Aug 22.
DOI:	10.1016/j.bbamem.2015.08.018
Abstrakt:	Apolipoproteins are essential human proteins for lipid metabolism. Together with phospholipids, they constitute lipoproteins, nm to μm sized particles responsible for transporting cholesterol and triglycerides throughout the body. To investigate specific protein-lipid interactions, we produced and characterized three single-Trp containing apolipoprotein C-III (ApoCIII) variants (W42 (W54F/W65F), W54 (W42F/W65F), W65 (W42F/W54F)). Upon binding to phospholipid vesicles, wild-type ApoCIII adopts an α-helical conformation (50% helicity) as determined by circular dichroism spectroscopy with an approximate apparent partition constant of 3×10(4) M(-1). Steady-state and time-resolved fluorescence measurements reveal distinct residue-specific behaviors with W54 experiencing the most hydrophobic environment followed by W42 and W65. Interestingly, time-resolved anisotropy measurements show a converse trend for relative Trp mobility with position 54 being the least immobile. To determine the relative insertion depths of W42, W54, and W65 in the bilayer, fluorescence quenching experiments were performed using three different brominated lipids. W65 had a clear preference for residing near the headgroup while W54 and W42 sample the range of depths ~8-11 Å from the bilayer center. On average, W54 is slightly more embedded than W42. Based on Trp spectral differences between ApoCIII binding to phospholipid vesicles and sodium dodecyl sulfate micelles, we suggest that ApoCIII adopts an alternate helical conformation on the bilayer which could have functional implications. (Published by Elsevier B.V.)
Databáze:	MEDLINE
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