Effect of pH on the Ultraviolet Optical Rotatory Dispersion and Circular Dichroism of Lysozyme<xref ref-type="fn" rid="fn1"><sup>*</sup></xref>

Abstract

In order to clarify the state of the tyrosyl and tryptophyl residues of hen and duck egg-white lysozymes [mucopeptide N-acetylmuramyl-hydrolase, EG 3.2.1.17], ultraviolet optical rotatory dispersion (ORD**) and circular dichroism (CD)** were measured. The CD spectrum of both lysozymes in a neutral solution had a positive band at 288 mμ and a negative band at 268 mμ. At alkaline pH, the maximum of the band at 288 mμ was shifted to 298 mμt. At pH above pH 12, however, the CD band at 298 mμ decreased with time and finally disappeared. Spectrophotometric titration of the tyrosyl groups and ORD measurement have shown that lysozyme is denatured on exposure to pH above 12 and one and two tyrosyl residues become titratable for hen and duck lysozyme, respectively. Therefore, it is suggested that the CD band at 298 mμ arises from ionized tyrosyl residues which locate on the surface of the lysozyme molecule and are dependent on the protein conformation. On the other hand, a positive CD band was also observed around 250 mμ at alkaline pH. This band did not disappear on alkali denatu-ration. An approximately linear relation was obtained between the values of molecular ellipticity at the maximum of the CD band around 250 mμ and the number of ionized tyrosyl residues. Therefore, the CD band around 250 mμ depends mainly on the number of ionized tyrosyl residues and involves little conformational contribution. However, contribution of other side chains to the optical activity around 250 mμ must be also taken into account.