Benzobactins are natural bacterial products that have special biological activity due to a complex consisting of two ring structures. The bacterial genes responsible for the formation of the compound were previously unclear. Now, scientists at the Max Planck Institute for Terrestrial Microbiology have deciphered their biology through extensive genetic research. Their research facilitates the discovery of many natural compounds previously unknown to prescription drug treatments.
Microorganisms in their natural environment are often exposed to changing environmental conditions that require several responses to survive. The most efficient of these is their ability to produce a wide variety of natural products with diverse chemical structures and functions.
Benzobactins – potent but rare
Benzoxazoline is a rare natural compound that imparts extraordinary biological activities to natural products. It is, for example, the essential part of lidamycin, an antitumor antibiotic and one of the most cytotoxic compounds to date. The reason for this ability is the fact that benzoxazolines have two rings, a structure that allows them to interact with protein as well as with DNA. However, tracking down the producers of this rare substance in nature is like looking for a needle in a haystack.
In order to exploit new natural compounds of pharmaceutical value, such as antibiotics, tumor suppressors or immunosuppressants, it is necessary to know the responsible genes, or more precisely, the biosynthetic gene clusters (BGCs). BGCs are locally assembled combinations of two or more genes that together encode the production of a particular group of enzymes – and thus the corresponding natural products produced by these enzymes.
To date, the benzoxazoline biosynthetic gene cassette has remained elusive, which has hampered expansion of the repertoire of benzoxazoline-containing bioactive compounds. More specifically, the final benzoxazoline formation step was not clear. A team of Max Planck scientists led by Dr. Yi-Ming-Shi and Prof. Dr. Helge Bode has now succeeded in biocharacterizing the benzoxazoline pathway. During biosynthesis, the pathway obviously “borrows” an intermediate from the so-called phenazine pathway, which is responsible for producing another natural product. Most importantly, the researchers identified the enzyme responsible for the final step, degradation toward benzoxazolines.
Use of the enzyme as a probe for natural materials
“Knowing the identity of the enzyme, we used it as a probe,” explains doctoral student Jan Krams, co-first author of the study that was also funded by the LOEWE Center for Transitional Biodiversity Genomics (LOEWE TBG) and the European Research Council. Closely related biosynthesis of benzoxazoline-containing natural products, called benzobactins.” According to the scientists, the most surprising aspect was the wide distribution of these genes in other bacteria. “These pathways have been found in remarkably taxonomically and ecologically diverse bacteria ranging from land to ocean, as well as plant pathogens and biocontrol microbes. Their wide distribution indicates that these molecules have an important ecological function on producers,” Yi-Ming Shi notes. First author of the study.
Professor Helge Bode, Head of the Department “Natural Products in Organic Reactions” at the Max Planck Institute for Terrestrial Microbiology in Marburg adds: “Our findings reveal the enormous biosynthetic potential of the wide-ranging biosynthetic gene pool of benzobactin. Now, we have to figure out its function environment and whether we can use them as antibiotics or other medicines.”
Crystallization and structural analysis of the key enzyme were performed in collaboration with Dr Laura Cheek and Professor Gert Pang of SYNMIKRO, Philipp University Marburg. Enzyme kinetics was analyzed by Nicole Bakzia and the team from our Primary Metabolomics and Small Molecular Mass Spectrometry Facility. The genome sequencing was performed by Professor Jorn Kalinowski and a team from the University of Bielefeld.