Details

Autor(en) / Beteiligte

• Torta, Federico
• Elviri, Lisa
• Careri, Maria
• Mangia, Alessandro
• Cavazzini, Davide
• Rossi, Gian Luigi

Titel

Mass spectrometry and hydrogen/deuterium exchange measurements of alcohol-induced structural changes in cellular retinol-binding protein type I

Ist Teil von

• Rapid communications in mass spectrometry, 2008-02, Vol.22 (3), p.330-336

Ort / Verlag

Chichester, UK: John Wiley & Sons, Ltd

Erscheinungsjahr

2008

Quelle

MEDLINE

Beschreibungen/Notizen

• To bind and release its ligand, cellular retinol‐binding protein type I (CRBP) needs to undergo conformational and dynamic changes to connect the inner, solvent‐shielded cavity, where retinol is found to bind, and the outside medium. Retinol dissociation in vitro is favoured by water/alcohol mixtures whose moderately low dielectric constants mimic a property characteristic of the membrane microenvironment where this process occurs in vivo. Apo‐ and holo‐CRBP, in either water/methanol or water/trifluoroethanol (TFE) mixtures, were analyzed at equilibrium by electrospray ionization with orthogonal quadrupole time‐of‐flight mass spectrometry (ESI‐Q‐TOFMS) to identify the alcohol‐induced species. The questions were asked whether the presence of alcohols affects protein dynamics, as reflected by hydrogen/deuterium (H/D) exchange monitored by continuous‐labelling experiments, and to which extent retinol dissociation influences the process. With increasing methanol, at pH near neutrality, apo‐CRBP exhibits a progressively more compact conformation, resulting in reduced H/D exchange with respect to the native protein in water. Retinol dissociation from the holo‐protein did not promote hydrogen replacement. Similarly, in the presence of the low TFE concentration sufficient to cause retinol dissociation, the hydrogen exchange of the resulting apo‐protein was not exalted. However, in contrast with the alkanol, higher TFE concentrations induced a transition of apo‐CRBP to a new α‐helix conformation capable of exchanging all available hydrogen atoms. Copyright © 2008 John Wiley & Sons, Ltd.