Identification of antibacterial peptides from endophytic microbiome
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Endophytes, microorganisms living inside plant tissues, are promising producers of lead compounds for the pharmaceutical industry. However, the majority of endophytes are unculturable and therefore inaccessible for functional studies. To evaluate genetic resources of endophytes, we analyzed the biodiversity of fungal microbiome of black crowberry (Empetrum nigrum L.) by next-generation sequencing and found that it consists mainly of unknown taxa. We then separated the host and the endophyte genomes and constructed a fosmid expression library from the endophytic DNA. This library was screened for antibacterial activity against Staphylococcus aureus. A unique antibacterial clone was selected for further analysis, and a gene En-AP1 was identified with no similarity to known sequences. The expressed, folded protein En-AP1 was not active against S. aureus, while tryptic digests exhibited antimicrobial activity. Seven out of twelve synthesized peptides, predicted antibacterial in silico, exhibited in vitro activity towards both S. aureus and Escherichia coli. We propose that the En-AP1 protein is degraded in the library host E. coli and antimicrobial fragments are released from the cell, explaining the in vitro antibacterial activity of the clone. This is the first report of a novel gene expressed in vitro derived from an endophytic microbiome, demonstrating the potential of finding novel genes and compounds from unculturable endophytes.
KeywordsAntimicrobial peptide Unculturable Endophyte Microbiome
The work was funded by the Marie Curie Industry-Academia Partnership and Pathways (IAPP) of the EU 7th Framework Programme, New Antimicrobials (NAM) Project (PIAP-GA-2008-218191) and PIIF-GA-2008-220253.
MVT, PP and AMP designed the study. MVT and PP performed molecular lab experiments. AMP, LR, AT, HHS, and HHK provided reagents, materials and scientific input to the study. MK, HH, SK, and SM performed HPLC and MS experiments. MVT analyzed experiments and prepared figures. MVT, PP, and AMP wrote the manuscript. All authors revised the manuscript. All authors read and approved the final manuscript.
Compliance with ethical standards
Conflict of Interest
The authors declare that they have no conflict of interest.
This article does not contain any studies with human participants or animals performed by any of the authors.
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