REPTILE LYSOZYME - THE COMPLETE AMINO-ACID-SEQUENCE OF SOFT-SHELLED TURTLE LYSOZYME AND ITS ACTIVITY

Soft-shelled turtle egg-white lysozyme was purified and sequenced. Lys ozyme,vas reduced and carboxymethylated to fragment it with trypsin, v s protease and CNBr. The peptides yielded were purified by RP-HPLC and sequenced. Every trypsin peptide was overlapped by V8 protease peptid es and CNBr fragment. The amino acid sequence was compared with other lysozymes. This lysozyme has an extra Gig residue at N-terminus, which was found in pheasant lysozyme. Further, this lysozyme has an inserti on of a Gly residue between 47 and 48 residues when compared with chic ken lysozyme, as found in human lysozyme, therefore it proved that thi s lysozyme has the largest number of amino acids (131 aa) in chicken t ype lysozymes. The amino acid substitutions were found at subsites E a nd F. Namely Phe34, Arg45, Thr47, and Arg114 were replaced by His, Tyr , Arg, and Tyr, respectively. The time course using N-acetylglucosamin e pentamer as a substrate showed a reduction of the rate constant for glycosidic cleavage and increase of binding free energy for subsites E and F, which proved the contribution of amino acids mentioned above f or substrate binding at subsites E and F.