Bioengineering of Lantibiotics: The Use of Lantibiotic Biosynthetic Enzymes for Biotechnology

Lantibiotics are ribosomally-synthesized antibacterial peptides that undergo extensive posttranslational modifications (PTM). These peptides hold great potential to treat bacterial infections caused by multi-drug resistant pathogens. A constraint in such compounds is the limited chemical-tools available for the enhancement of their bioactivity. An alternative is to use biosynthetic approaches by utilizing lantibiotic biosynthetic enzymes such as the ones present in microbisporicin biosynthesis. Apart from the PTMs present in all lantibiotics, microbisporicin contains an S-[(Z)-2-aminovinyl]-D-cysteine (MibD), a 5-chlorotryptophan (MibH) and a 3,4-dihydroxyproline (MibO). Bioactivity assays revealed that microbisporicin was 15 times more potent than the widely used lantibiotic nisin. Due to its structural similarity with nisin, a hypothesis explaining microbisporicin’s bioactivity is the presence of these PTMs. To exploit the potential use of MibD, MibH and MibO as tools to modulate the bioactivity of other lantibiotics it is essential to biochemically characterize them. Using recombinant technology in Escherichia coli and in vitro assays we reconstituted the enzymatic activity of MibD. When subjected to UV-visible spectroscopy MibD showed a spectrum characteristic of oxidized flavoproteins. MibD did not require the N-terminal leader sequence of its substrate to oxidative decarboxylate Cys24 as determined by mass spectrometry. Sequence similarity analysis provided further evidence to assign MibD to the family of homo-oligomeric flavin dependent Cys decarboxylases. Future efforts include the in vitro reconstitution of MibH and MibO as well as determination of substrate promiscuity of these enzymes. Such understanding will enable us to exploit the enzyme’s biotechnological potential of creating lantibiotic analogs carrying these modifications