Intramolecular long-range electron transfer (LRET) in hen egg-white lysozyme (HEWL) accompanying Trp --> TyrO radical transformation was investigated in aqueous solution by pulse radiolysis as a function of pH (5.2-7.4) and temperature (283-328K). The reaction was induced by highly selective oxidation of Trp with N-3(.) radicals under low concentration of the reactants but at a high HEWL/N-3(.) molar ratio, so that more than 99% of the oxidized protein molecules contained only a single tryptophyl radical. Synchronous decay of Trp(.) and build-up of TyrO(.) conformed satisfactorily to first-order kinetics, indicating that LRET involved either one or more Trp(.)/Tyr redox pairs characterized by similar rate constants. The rate constant of LRET, k(5), increased monotonously with decreasing pH showing the following characteristics: (i) in the pH range 7.4-5.2 the plot of k(5) against pH was sigmoidal in shape, reflecting protonation of Glu35 (pK(a) approximate to 6) and pointing to involvement of conformational control of the kinetics of LRET, (ii) below pH5.2 a sharp increase in k(5) was observed due to the protonation of Trp(.) to form TrpH(.+), which is known to oxidize tyrosine faster than does Trp(.). Arrhenius plots of the temperature-dependence of k(5) showed that the activation energy of LRET varies both with temperature and the protonation state of the enzyme. The activation energies are in the range 7.6-56.0 kJ mol(-1) and are similar to those for activation of amide hydrogen exchange in native HEWL below its denaturation temperature. Selective oxidation by ozone of the Trp62 indole side-chain in HEWL to N'-formylkynurenine (NFKyn62-HEWL) caused a large drop in the initial yield of Trp(.) radicals, G(Trp(.))(i). This was accompanied by a relatively small decrease in k(5) but selective oxidation by ozone had a pronounced effect on its temperature-dependence. Taken together these observations indicate that of the six tryptophans present in HEWL Trp62 contributes about 50% to the yield of the observed LRET. In the enzyme-inhibitor complex, HEWL(GlcNAc)(3), where Trp62 and Trp63 are completely shielded from the solvent by the bound triacetylchitotriose, G(Trp(.))(i) was lower than in NFKyn62-HEWL, and both the kinetic and energetic characteristics of LRET, observed at pH5.2, were again somewhat different than in HEWL alone. Considering known solvent accessibilities of tryptophans in the complex, the observed LRET process in HEWL(GlcNAc)(3) was assigned to Trp123. Theoretical evaluation of the electronic coupling for the dominant LRET pathways between all the potential Trp(.)/Tyr redox couples in HEWL, with help of the PATHWAYS model, enabled Trp62/Tyr53, Trp63/Tyr53 and Trp123/Tyr23 to be identified as the pairs involved in the experimentally observed electron transfer.