In the cross-linking reaction of lysozyme between Leu129 (α-COO-) and Lys13 (ε-NH3+ using imidazole and 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide hydrochloride (EDC), a side reaction of the peptide bond inversion from α to β between A and Gly102 was greatly reduced by addition of β-(1,4)-linked trimer of N-acetyl-D-glucosamine [(NAG)3] When methylamine or 2-hydroxyethylamine was further added, the extent of the cross-link formation was decreased and the derivative where the α-carboxyl group of Leu129 was modified with the amine was newly obtained. On the other hand, when ammonia was added, the β-carboxyl group of Asp119 instead of the α-carboxyl group was mainly amidated. From these results, the presence of a salt bridge between Asp119 and Arg125 besides that between Lys13 and Leu129 is proposed. Enzymatic activities of the derivatives prepared here indicated that the modification of the α-carboxyl group reduced the activity to ̃ 90% of that of native lysozyme. Des-Leu129 lysozyme, which lacks Leu129 also showed ̃ 90% of the activity of native lysozyme. Therefore, the salt bridge between Lys13 and Leu129 may play some role in maintaining the active conformation of lysozyine.
All Science Journal Classification (ASJC) codes
- Molecular Biology