Leinamycin (LNM), migrastatin (MGS), dorrigocin (DGN), and lactimidomycin (LTM) encompass a broad spectrum of biological activities. This application proposes (1) to study the biochemistry and genetics of LNM, MGS/DGN, and LTM production as models for """"""""AT-less"""""""" type I polyketide synthase (PKS)- catalyzed polyketide biosynthesis, (2) to develop new combinatorial biosynthesis strategies for engineering polyketide structural diversity, and (3) to manipulate LNM, MGS/DGN, and LTM biosyntheses for the production of biologically active compounds. The hypotheses are: (1) """"""""AT-less"""""""" type I PKSs represent an unprecedented PKS structure, the studies of which will reveal new insights into the molecular mechanism of PKS catalysis; (2) """"""""AT-less"""""""" type I PKS provides new opportunities for PKS engineering, methods and strategies based on which will further expand the number and diversity of polyketide products accessibly by combinatorial biosynthesis; (3) LNM, MGS/DGN, and LTM biosyntheses are unprecedented, the characterization of which will uncover new chemistry for polyketide biosynthesis; and (4) LNM, MGS/DGN, and LTM are excellent starting molecular scaffolds for combinatorial biosynthesis, derivation of which could lead to the discovery of therapeutically useful agents. The outcome of the proposed studies will (1) expand the repertoire of PKS genes for combinatorial biosynthesis; (2) provide new strategies and methods for combinatorial biosynthesis employing """"""""AT-less"""""""" type I PKS; and (3) potentially lead to the production of LNM, MGS/DGN, and LTM analogs as novel therapeutic agents.
The specific aims for this grant period are: (1) functional analysis of the Inm biosynthetic gene cluster by targeted gene inactivation in vivo in S. atroolivaceus and by biochemical characterization of the LNM hybrid nonribosomal peptide synthetase (NRPS)-""""""""AT-less"""""""" type I PKS megasynthetase in vitro; (2) functional analysis of the mgs/dgn biosynthetic gene cluster by targeted gene inactivation in vivo in S. platensis and by biochemical characterization of the MGS/DGN """"""""AT-less"""""""" type I PKS in vitro; (3) functional analysis of the Itm biosynthetic gene cluster by targeted gene inactivation in vivo in S. amphibiosporus and by biochemical characterization of the LTM """"""""AT-less"""""""" type I PKS in vitro; (4) development of combinatorial biosynthesis strategies and methods based on the LNM, MGS/DGN, LTM biosynthetic machinery for novel polyketide production.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
1R01CA106150-01
Application #
6710530
Study Section
Special Emphasis Panel (ZRG1-SSS-B (01))
Program Officer
Fu, Yali
Project Start
2004-05-01
Project End
2009-04-30
Budget Start
2004-05-01
Budget End
2005-04-30
Support Year
1
Fiscal Year
2004
Total Cost
$296,617
Indirect Cost
Name
University of Wisconsin Madison
Department
Type
Schools of Pharmacy
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Teijaro, Christiana N; Adhikari, Ajeeth; Shen, Ben (2018) Challenges and opportunities for natural product discovery, production, and engineering in native producers versus heterologous hosts. J Ind Microbiol Biotechnol :
Kwong, Thomas; Ma, Ming; Pan, Guohui et al. (2018) P450-Catalyzed Tailoring Steps in Leinamycin Biosynthesis Featuring Regio- and Stereoselective Hydroxylations and Substrate Promiscuities. Biochemistry 57:5005-5013
Rudolf, Jeffrey D; Chang, Chin-Yuan; Ma, Ming et al. (2017) Cytochromes P450 for natural product biosynthesis in Streptomyces: sequence, structure, and function. Nat Prod Rep 34:1141-1172
Pan, Guohui; Xu, Zhengren; Guo, Zhikai et al. (2017) Discovery of the leinamycin family of natural products by mining actinobacterial genomes. Proc Natl Acad Sci U S A 114:E11131-E11140
Zhang, Bo; Xu, Zhengren; Teng, Qihui et al. (2017) A Long-Range Acting Dehydratase Domain as the Missing Link for C17-Dehydration in Iso-Migrastatin Biosynthesis. Angew Chem Int Ed Engl 56:7247-7251
Huang, Yong; Yang, Dong; Pan, Guohui et al. (2016) Characterization of LnmO as a pathway-specific Crp/Fnr-type positive regulator for leinamycin biosynthesis in Streptomyces atroolivaceus and its application for titer improvement. Appl Microbiol Biotechnol 100:10555-10562
Smanski, Michael J; Zhou, Hui; Claesen, Jan et al. (2016) Synthetic biology to access and expand nature's chemical diversity. Nat Rev Microbiol 14:135-49
Huang, Yong; Tang, Gong-Li; Pan, Guohui et al. (2016) Characterization of the Ketosynthase and Acyl Carrier Protein Domains at the LnmI Nonribosomal Peptide Synthetase-Polyketide Synthase Interface for Leinamycin Biosynthesis. Org Lett 18:4288-91
Zhang, Bo; Yang, Dong; Yan, Yijun et al. (2016) Overproduction of lactimidomycin by cross-overexpression of genes encoding Streptomyces antibiotic regulatory proteins. Appl Microbiol Biotechnol 100:2267-77
Yang, Zhilong; Cao, Shuai; Martens, Craig A et al. (2015) Deciphering poxvirus gene expression by RNA sequencing and ribosome profiling. J Virol 89:6874-86

Showing the most recent 10 out of 46 publications