Abstract:
Cyclic lipopeptides like surfactin, iturin, fengycine etc. are a group of natural products with a wide range of promising properties, including antibiotic and surface activity. They usually consist of a peptide ring and a lipid moiety. In times of emerging antibiotic-resistant bacteria lipopeptides could offer new structures for the solution of this problem. Furthermore, due to their surface activity they could also be used as environment friendly oil dispersant.
Cyclic lipopeptides are produced by nonribosomal-peptide synthetases (NRPS) or occasionally in combination with polyketide synthases (PKS). NRPS and PKS show a modular structure which is reflected in their genetic organization. This modular structure genetic organization could be utilized to create new derivatives of cyclic lipopeptides. Modules could be deleted, added or replaced by others to create new cyclic lipopeptides derivatives.
We are using the surfactin NRPS in Bacillus subtilis 168 to create novel surfactin derivatives. The peptide ring of surfactin consists of seven amino acids and its lipid chain ranges in size from C12 to C16. Our focus was put on the modification of the peptidic portion. The amino acid composition and order of surfactin is glutamic acid, leucine, D-leucine, valine, aspartic acid, D-leucine and followed by leucine. In our work we aimed at the modification of the stereochemistry of surfactin, the charged amino acid at position five, the leucine at position seven, the lactam ring formation and the creation of novel lipopeptides utilizing COM-domains. The charged amino acids should be deleted or replaced to increase the surfactin stability in sea water. The stereochemistry of surfactin was modified by introducing an E-domain in module 5 and by deleting the E-domain in module 6. The charged amino acid at position 5 was removed by using three different approaches and resulted in the production of [ΔAsp5]surfactin and [ΔAsp5ΔLeu7]surfactin derivatives. Leucin at position seven was removed in three different ways and unveiled different mechanisms of how [ΔLeu7]surfactin derivatives could be formed. The production of lactamo-surfactin was not achieved, but we were able to determine factors that could contribute to the success of this project in future. A further way to generate novel lipopeptides could be the utilization of COM-domains in order to create new cross communications between different NRPS subunits. We were not able to introduce cross communication between the surfactin and the fengycin gene cluster. However, based on the results of this study we developed an optimized concept to achieve this in future.
