Structural Dynamics of Soluble Chloride Intracellular Channel Protein CLIC1 Examined by Amide Hydrogen-Deuterium Exchange Mass Spectrometry

摘要

ABSTRACT: Chloride intracellular channel protein 1 (CLIC1) functions as an anion channel in plasma andnnuclear membranes when its soluble monomeric form converts to an integral-membrane form. Thentransmembrane region of CLIC1 is located in its thioredoxin-like domain 1, but the mechanism wherebynthe protein converts to its membrane conformation has yet to be determined. Since channel formation innmembranes is enhanced at low pH (5 to 5.5), a condition that is found at the surface of membranes, thenstructural dynamics of soluble CLIC1 was studied at pH 7 and at pH 5.5 in the absence of membranes bynamide hydrogen-deuterium exchange mass spectrometry (DXMS). Rapid hydrogen exchange data indicatenthat CLIC1 displays a similar core structure at these pH values. Domain 1 is less stable than the all-helicalndomain 2, and, while the structure of domain 1 remains intact, its conformational flexibility is furthernincreased in an acidic environment (pH 5.5). In the absence of membrane, an acidic environment appears tonprime the solution structure of CLIC1 by destabilizing domain 1 in order to lower the activation energynbarrier for its conversion to themembrane-insertion conformation. The significantly enhancedH/D-exchangenrates at pH 5.5 displayed by two segments (peptides 11-31 and 68-82) could be due to the protonation ofnacidic residues in salt bridges. One of these segments (peptide 11-31) includes part of the transmembranenregion which, in the solution structure, consists of helix R1. This helix is intrinsically stable and is most likelynretained in the membrane conformation. Strand β2, another element of the transmembrane region, displays anpropensity to form a helical structure and has putative N- and C-capping motifs, suggesting that it too mostnlikely forms a helix in a lipid bilayer.