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Hydrogen–Deuterium Exchange in Bovine Serum Albumin Protein Monitored by Fourier Transform Infrared Spectroscopy, Part II: Kinetic Studies

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Abstract:

The set of infrared spectra recorded at different levels of hydrogen–deuterium (H/D) exchange as a function of time were processed using spectral decomposition. The most precise information about H/D exchange of the NH groups of the protein backbone was retrieved by observing the intensity change of the nearest C=O stretching vibration. The H/D exchange at the protein backbone begins with NH groups bonded to C=O with a characteristic frequency of 1683 cm−1. These amide groups were initially free and are the first to accept H-bonds from water during the hydration process. The NH groups, which are connected to C=O groups with a characteristic band at 1657 cm−1, exchange at a slower rate. For both populations the pairs of comparable exchange rates were calculated with rate constants of 10−3 min−1, 0.014 min−1, 0.0046 min−1, and 0.09 min−1. The appearance of two different exchange rates for each population is a consequence of the distinct exposure of particular molecular groups to the solvent. Two additional bands sensitive to exchange are attributed to NH bending modes in the side chains and are located at 1610 cm−1 and 1585 cm−1. These NH groups undergo H/D exchange at the beginning of the exchange with exchange rates of 0.019 min−1 and 0.17 min−1, respectively. The calculated exchange rate for hydrating water molecules is 0.037 min−1. The distribution of water's exchange rate is extremely broad and covers almost the entire interval of the time-dependent experiment. From the efficiency of the exchange (96%) it is evident that some parts of the protein are completely forbidden to water molecules.

Keywords: BOVINE SERUM ALBUMIN; CALCULATION OF RATE CONSTANTS; CLASSES OF EXCHANGE; FOURIER TRANSFORM INFRARED SPECTROSCOPY; FT-IR SPECTROSCOPY; SPECTRAL DECOMPOSITION; TIME-DEPENDENT H/D EXCHANGE

Document Type: Research Article

DOI: http://dx.doi.org/10.1366/000370205774783287

Affiliations: 1: National Institute of Chemistry, Hajdrihova 19, SI-1000 Ljubljana, Slovenia 2: Département de Recherche Fondamentale sur la Matiére Condensée, SI3M/PCM, CEA Grenoble, 17 rue des Martyrs, F 38054 Grenoble Cedex 9, France

Publication date: November 1, 2005

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