There is a critical need for new pharmaceutical agents against a wide variety of old and emerging human diseases, such as AIDS, antibiotic-resistant pathogens such as Mycobacterium tuberculosis, Staphylococcus aureus, and Pseudomonas aeruginosa, fungal diseases, and many types of cancer. However, the rate at which new types of compounds are being discovered has slowed. Natural products (chemicals made by living organisms such as bacteria and fungi) have historically been the source of most of our medicines. The natural world provides multiple examples of novel chemical classes with medically relevant biological activities. One is exemplified by the cyclic peptide toxins of fungi in the genus Amanita. These compounds have several unique and desirable properties among known natural products that make them promising scaffolds on which to design new therapeutic compounds for improving human health. For example, they are the smallest known peptides synthesized on ribosomes, and some of them already are known to have strong and unique biological targets. Methods used in the proposed studies include molecular genetics, next-generation deep genome sequencing, bioinformatics, biochemistry, chemistry, and bioassays against target organisms and mammalian cancer lines.
The specific aims of this proposal are, first, to identify all of the steps in the biosynthesis of the amatoxins and phallotoxins, which are predicted to include proteolytic processing, cyclization, hydroxylation, synthesis of a unique amino acid crossbridge (trypathionine), and, in the case of the phallotoxins, conversion of one amino acid from the L to the unusual D form.
The second aim i s to study the biosynthesis of the amatoxins in Galerina, a mushroom unrelated to Amanita that makes the same compounds. The work on Amanita and Galerina will complement each other in several ways, leading to a more definitive description of the biosynthetic pathway.
The third aim i s to express the toxin biosynthetic pathway in another host organism that is more genetically tractable than Amanita itself. This will establish an experimental platform to dissect the pathway in detail, and to modify it in beneficial ways.
The fourth aim i s to use this experimental system to synthesize in vivo new cyclic peptides based on the Amanita toxin scaffold. There are more than 3 million possible permutations of amanitin, and this library of novel compounds is predicted to be rich in chemicals of potential pharmaceutical importance for treatment of bacterial and fungal diseases, and cancer.

Public Health Relevance

Most of our current antibiotics and anticancer compounds come from natural sources such as bacteria, fungi, and plants. Certain mushrooms in the genus Amanita produce bioactive cyclic peptides with a range of known or potential uses in biomedical research and clinical medicine. This proposal describes basic research on understanding how this unique class of natural products is biosynthesized, with the goal of producing families of new compounds of medical utility.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM088274-04
Application #
8642189
Study Section
Synthetic and Biological Chemistry B Study Section (SBCB)
Program Officer
Gerratana, Barbara
Project Start
2011-06-15
Project End
2015-03-31
Budget Start
2014-04-01
Budget End
2015-03-31
Support Year
4
Fiscal Year
2014
Total Cost
$285,456
Indirect Cost
$97,656
Name
Michigan State University
Department
Other Basic Sciences
Type
Schools of Arts and Sciences
DUNS #
193247145
City
East Lansing
State
MI
Country
United States
Zip Code
48824
Riley, Robert; Salamov, Asaf A; Brown, Daren W et al. (2014) Extensive sampling of basidiomycete genomes demonstrates inadequacy of the white-rot/brown-rot paradigm for wood decay fungi. Proc Natl Acad Sci U S A 111:9923-8
Sgambelluri, R Michael; Epis, Sara; Sassera, Davide et al. (2014) Profiling of amatoxins and phallotoxins in the genus Lepiota by liquid chromatography combined with UV absorbance and mass spectrometry. Toxins (Basel) 6:2336-47
Luo, Hong; Hong, Sung-Yong; Sgambelluri, R Michael et al. (2014) Peptide macrocyclization catalyzed by a prolyl oligopeptidase involved in ?-amanitin biosynthesis. Chem Biol 21:1610-7
Luo, Hong; Hallen-Adams, Heather E; Scott-Craig, John S et al. (2012) Ribosomal biosynthesis of ?-amanitin in Galerina marginata. Fungal Genet Biol 49:123-9