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Appl Environ Microbiol. 2019 Aug 9. pii: AEM.01292-19. doi: 10.1128/AEM.01292-19. [Epub ahead of print]

The ADEP biosynthetic gene cluster in Streptomyces hawaiiensis NRRL 15010 reveals an accessory clpP gene as a novel antibiotic resistance factor.

Author information

1
Department of Microbial Bioactive Compounds, Interfaculty Institute of Microbiology and Infection Medicine, University of Tuebingen, Tuebingen, Germany.
2
M. G. DeGroote Institute for Infectious Disease Research, Department of Biochemistry and Biomedical Sciences, David Braley Centre for Antibiotic Discovery, McMaster University, Hamilton, Ontario, Canada.
3
Department of Applied Natural Products Genome Mining, Interfaculty Institute of Microbiology and Infection Medicine, University of Tuebingen, Tuebingen, Germany.
4
UNT System College of Pharmacy, University of North Texas Health Science Center, Fort Worth, Texas, USA.
5
Department of Microbial Bioactive Compounds, Interfaculty Institute of Microbiology and Infection Medicine, University of Tuebingen, Tuebingen, Germany heike.broetz-oesterhelt@uni-tuebingen.de.

Abstract

The increasing threat posed by multi-resistant bacterial pathogens necessitates the discovery of novel antibacterials with unprecedented modes of action. ADEP1, a natural compound produced by Streptomyces hawaiiensis NRRL 15010, is the prototype for a new class of acyldepsipeptide (ADEP) antibiotics. ADEP antibiotics deregulate the proteolytic core ClpP of the bacterial caseinolytic protease, thereby exhibiting potent antibacterial activity against Gram-positive bacteria, including multi-resistant pathogens. ADEP1 and derivatives, here collectively called ADEP, have been previously investigated for their antibiotic potency against different species, structure-activity relationship, and mechanism of action, however, knowledge on the biosynthesis of the natural compound and producer self-resistance has remained elusive. In this study, we identified and analyzed the ADEP biosynthetic gene cluster in S. hawaiiensis NRRL 15010, which comprises two NRPSs, genes necessary for the biosynthesis of (4S,2R)-4-methylproline, and a type II PKS for the assembly of highly reduced polyenes. Whilst no resistance factor could be identified within the gene cluster itself, we discovered an additional clpP homologous gene (named clpPADEP ) located further downstream of the biosynthetic genes, separated from the biosynthetic gene cluster by several transposable elements. Heterologous expression of ClpPADEP in three ADEP-sensitive Streptomyces species proved its role in conferring ADEP resistance, thus revealing a novel type of antibiotic resistance determinant.IMPORTANCEAntibiotic acyldepsipeptides (ADEPs) represent a promising new class of potent antibiotics and, at the same time, are valuable tools to study the molecular functioning of their target ClpP, the proteolytic core of the bacterial caseinolytic protease. We here present a straightforward purification procedure for ADEP1 that yields substantial amounts of the pure compound in a time- and cost-efficient manner, which is a prerequisite to conveniently study the antimicrobial effects of ADEP and the operating mode of bacterial ClpP machineries in diverse bacteria. Identification and characterization of the ADEP biosynthetic gene cluster in Streptomyces hawaiiensis NRRL 15010 enables future bioinformatics screenings for similar gene clusters and/or sub-clusters, to find novel natural compounds with specific sub-structures. Most strikingly, we identified a cluster-associated clpP homolog (named clpPADEP ) as ADEP resistance gene. ClpPADEP constitutes a novel bacterial resistance factor, alone necessary and sufficient to confer high-level ADEP resistance to Streptomyces across species.

PMID:
31399403
DOI:
10.1128/AEM.01292-19

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