Transglutaminase-mediated crosslinking of a host defence peptide derived from human apolipoprotein B and its effect...

Background Microbial transglutaminase (mTG) has been successfully used to produce site-specific protein conjugates derivatized at the level of Gln and/or Lys residues for different biotechnological applications. Here, a recombinant peptide identified in human apolipoprotein B sequence, named r(P)ApoB_L and endowed with antimicrobial activity, was studied as a possible acyl acceptor substrate of mTG with at least one of the six Lys residues present in its sequence. Methods The enzymatic crosslinking reaction was performed in vitro using N,N-dimethylcasein, substance P and bitter vetch (Vicia ervilia) seed proteins, well known acyl donor substrates in mTG-catalyzed reactions. Mass spectrometry analyses were performed for identifying the Lys residue(s) involved in the crosslinking reaction. Finally, bitter vetch protein-based antimicrobial films grafted with r(P)ApoB_L were prepared and, their biological activity evaluated. Results r(P)ApoB_L was able to be enzymatically modified by mTG. In particular, it was demonstrated the highly selective crosslinking of the peptide under study by mTG at level of Lys-18. Interestingly, the biological activity of the peptide when grafted into protein-based films was found to be lost following mTG-catalyzed crosslinking. Conclusions r(P)ApoB_L was shown to be an effective acyl acceptor substrate of mTG. The involvement of Lys-18 in the enzymatic reaction was demonstrated. In addition, films grafted with r(P)ApoB_L in the presence of mTG lost antimicrobial property. General significance A possible role of mTG as biotechnological tool to modulate the r(P)ApoB_L antimicrobial activity was hypothesized, and a potential use in food packaging of protein-based films grafted with r(P)ApoB_L was suggested.