New Pipeline to Develop Antimicrobial Lanthipeptides

Nature has provided us with lots of antimicrobials. However, given the rapid increase of antimicrobial resistance, there is a need for the development of new-to-nature antibiotics. In a study published yesterday in Nature Chemical Biology, a team of scientists developed a pipeline to create and screen large numbers of new lantibiotic peptides.

Lantibiotic peptides are modified after production by enzymes that link different amino acids within the peptide chain to form rings. “There are different amino acid sequences that make up the rings. We know that a selection of 12 natural lantibiotics all have different combinations,” explains Oscar Kuipers of the University of Groningen. “We synthesized DNA strands coding for the different rings and combined these genetic modules to form lantibiotic genes coding for random combinations of five rings.”

The next step was to screen the gene products for antimicrobial potential. To do this, they modified a technique developed for enzyme screening, based on micro-alginate beads. Inside these beads, bacteria can grow. The genes from the library were put into a producer strain, containing a red fluorescent marker.

“We diluted these bacteria so that we would end up with one or zero cells per bead. We also added a target strain that could be killed by the lantibiotics; those cells were used at a higher concentration of around 50 per bead,” Kuipers says.

The target cells carried a green fluorescent marker and produced a peptidase that activated the precursor lantibiotics secreted by the producer cells.

When the beads were incubated, the producer strain formed a single colony inside, secreting lantibiotics. These peptides could not leave the beads. The number of green colonies growing inside each bead was negatively correlated with the antimicrobial effect of the activated lanthipeptides.

“We found peptides with different antibiotic activities, although several peptides looked a bit like nisin or epidermin. It turned out that we had even recreated nisin in our gene library,” Kuipers says. The gene library that they created contained around 14,000 synthetic genes.

The experiment provided a lot of information about the structure–function relationship of the different rings in the lantibiotic peptides. This can now be used for a rational design approach to create new antibiotics.

Image: Alginate microbeads containing colonies of target cells (green) and producer cells (red). Where the producer cells indeed secrete active lantibiotics, the target cells have disappeared. Image courtesy of Steven Schmitt ETH Zürich.