N⁶-Cbz-L-lysine is an amino acid derivative, and its enzymatic hydrolysis characteristics are closely related to the type of enzyme used. Different enzymes exhibit distinct hydrolysis effects and produce different products due to their specificity, as detailed below:
Enzymatic hydrolysis characteristics: Trypsin can specifically recognize and cleave lysine residues. The lysine moiety in N⁶-Cbz-L-lysine can be recognized by trypsin, exposing it to the reaction interface and thereby undergoing cleavage. Additionally, Lys-C protease derived from Lysobacter enzymogenes specifically hydrolyzes the carboxyl terminal of lysine (except when linked to arginine). Therefore, N⁶-Cbz-L-lysine may also serve as a substrate for Lys-C protease and be hydrolyzed by it. Studies have also shown that carboxypeptidase Capa from Aspergillus niger has a certain hydrolytic capacity for substrates such as Cbz-Ala-Lys, with a preference for hydrolyzing substrates where the carboxyl terminal is a hydrophobic amino acid (e.g., leucine and lysine). Thus, N⁶-Cbz-L-lysine may potentially be hydrolyzed as its substrate.
Product analysis: When trypsin or Lys-C protease is used for enzymatic hydrolysis, cleavage occurs at the carboxyl terminal of N⁶-Cbz-L-lysine, potentially producing peptide segments or amino acid fragments containing a benzyloxycarbonyl (Cbz)-protected amino group. If the hydrolysis degree is low, oligopeptides containing N⁶-Cbz-L-lysine may be generated; if hydrolysis is complete, N⁶-Cbz-L-lysine monomers are obtained. When enzymes acting on the carboxyl terminal (such as carboxypeptidase Capa) are used, the carboxyl-terminal amino acids of N⁶-Cbz-L-lysine may be gradually removed. Upon complete hydrolysis, the final product may be Cbz-protected lysine. Under acidic conditions, the benzyl-protected ester group can be removed, releasing free lysine. Furthermore, according to relevant studies, N⁶-Cbz-L-lysine can undergo oxidation reactions under the action of certain microorganisms or enzymes to generate corresponding keto acids, which can then be reduced to produce Cbz-L-hydroxylysine derivatives.
