Purity of mushroom tyrosinase as a biocatalyst for biomaterial synthesis affects the stability of therapeutic peptides

Abstract

The formation of injectable implants in the presence of cells or solutes has previously been conceptualized to be based on the selectivity of bioorthogonal chemical reactions. As an alternative approach, hydrogel network synthesis by enzymatic reactions with a typically high inherent substrate specificity and low toxicity have been repeatedly proposed, e.g. using commercial mushroom tyrosinase (MTyr), which specifically catalyzes phenol oxidation. In this study, it should be explored whether MTyr is compatible with therapeutic peptides that may be delivered from such hydrogels in the future. Based on the specificity of MTyr to phenol residues, no modification of peptides lacking the amino acid tyrosine would be expected. One example of such peptides is gramicidin S (GS), a potent antimicrobial peptide. However, when GS was incubated with commercial MTyr, peptide degradation occurred as observed by HPLC analysis. Several fragments of the peptide were detected by MALDI-TOF. Contamination of MTyr with peptidases was proven as the source of undesired peptide cleavage, which needs to be considered when preparing enzymatically crosslinked hydrogels for biomedical applications.