Please use this identifier to cite or link to this item: http://hdl.handle.net/10397/9984
Title: Membrane-inserted conformation of transmembrane domain 4 of divalent-metal transporter
Authors: Li, H
Li, F
Sun, H
Qian, ZM
Keywords: Circular dichroism (CD)
Detergent
Divalent-metal transporter 1 (DMT1)
NMR
Phospholipid vesicles
Secondary structure
Issue Date: 2003
Publisher: Portland Press
Source: Biochemical journal, 2003, v. 372, no. 3, p. 757-766 How to cite?
Journal: Biochemical journal 
Abstract: Divalent-metal transporter 1 (DMT1) is involved in the intestinal iron absorption and in iron transport in the transferrin cycle, It transports metal ions at low pH (≈ 5.5), but not at high pH (7.4), and the transport is a proton-coupled process. Previously it has been shown that transmembrane domain 4 (TM4) is crucial for the function of this protein. Here we provide the first direct experimental evidence for secondary-structural features and membrane insertions of a 24-residue peptide, corresponding to TM4 of DMT1 (DMT1-TM4), in various membrane-mimicking environments by the combined use of CD and NMR spectroscopies. The peptide mainly adopts an α-helical structure in trifluoroethanol, SDS and dodecylphosphocholine micelles, and dimyristoyl phosphatidylcholine and dimyristoyl phosphatidylglycerol small unilamellar vesicles. It has been demonstrated from both Hα secondary shifts and nuclear-Overhauser-enhancement (NOE) connectivities that the peptide is well folded into an a-helix from Val8 to Lys23 in SDS micelles at pH 4.0, whereas the N-terminus is highly flexible. The α-helical content estimated from NMR data is in agreement with that extracted from CD simulations. The highest helicity was observed in the anionic phospholipids {1,2-dimyristoyl-sn-glycero-3-[phosphorac-(1-glycerol)]}, indicating that electrostatic attraction is important for peptide binding and insertion into the membranes. The secondary-structural transition of the peptide occurred at pH 4.3 in the 2,2,2-trifluoroethanol (TFE) water mixed solvent, whereas at a higher pH value (5.6) in SDS micelles, DMT1-TM4 exhibited a more stable structure in SDS micelles than that in TFE in terms of changing the pH and temperature. PAGE did not show high-molecular-mass aggregates in SDS micelles. The position of the peptide relative to SDS micelles was probed by the effects of 5- and 16-doxylstearic acids on the intensities of the peptide proton resonances. The results showed that the majority of the peptide is inserted into the hydrophobic interior of SDS micelles, whereas the C-terminal residues are surface-exposed. The ability of DMT1-TM4 to assume transmembrane features may be crucial for its biological function in vivo.
URI: http://hdl.handle.net/10397/9984
ISSN: 0264-6021
EISSN: 1470-8728
DOI: 10.1042/BJ20030075
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