Analysis of catalytic properties of hen egg white lysozyme during renaturation from denatured and reduced material.

Catalytic properties of hen egg white lysozyme were analyzed during the renaturation of the enzyme from completely reduced and denatured material. The formation of intermediate folding products and the generation of native lysozyme was monitored by acetic acid/urea electrophoresis. The results showed that during the beginning of renaturation almost all reduced and denatured lysozyme is converted to forms possessing lower compactness than native lysozyme, probably as a result of formation of only one or two disulfide bonds. Kinetic analysis of lysozyme during renaturation showed that the generation of lysozyme with four disulfide bonds was not necessarily equivalent to the formation lysozyme with native-like catalytic properties. It appeared that the formation rate of the structures of the structures of the substrate binding site and of the catalytic site were limited by the generation of four disulfide bonds containing lysozyme. The catalytic properties of intermediate folding products made it evident that the final structures of the substrate binding site and of the catalytic site were formed after the generation of all disulfide bonds.