The role of protein structure in the reactivity of the four disulfide (S-S) bridges of lysozyme was studied using Raman spectroscopy and molecular modelling. The experimental kinetics of S-S bridge reduction by tris-2-carboxyethyl phosphine (TCEP) was obtained by monitoring the protein S-S Raman bands. The kinetics are heterogeneous and were fitted using two apparent reaction rate constants. Kinetic measurements performed at different pH values indicate only moderate charge effects. The two intrinsic reaction rate constants derived for the neutral TCEP species were 0.45 and 0.052 mol(-1) s(-1), respectively. The molecular dynamics simulation of the reactants encounter shows that the accessibility of the lysozyme S-S bridges by TCEP decreases in the following order: cys30-cys115 > cys6-cys127 > cys64-cys80 > cys76-cys94. This simulation also illustrates the reaction mechanism which consists of a local unfolding followed by the reduction of the exposed S-S bridge. The Gibbs free energy for local unfolding was evaluated by comparing the actual reaction rate constant with that of a model system containing a fully exposed S-S bridge (oxidized glutathione). These values corresponding to the fast- and slow-reaction rate-constants were 8.5 and 13.8 kJ mol(-1), respectively. On the other hand, Raman measurements, as well as the molecular dynamics simulations, strongly suggest that the protein global unfolding following S-S bridge cleavage has only limited effects in stabilizing the reaction products.