Many biologically active natural products derive their activity from the sugar components of their structures. Changing the structures of these sugars can have a profound impact on the biological activity, selectivity, and pharmacokinetic properties of the parent compounds. This observation has fueled the development of methods to derivatize natural products with diverse sugar moieties by exploiting the sugar biosynthetic machinery. Fully realizing the potential of such an approach relies on the discovery of new sugar biosynthetic pathways, and also requires a thorough understanding of the biosynthetic pathway of each target sugar including detailed mechanistic knowledge of the key enzymes. With these goals in mind, we have produced notable results in work funded by previous grant. As a result of these studies, we have identified four key areas that warrant further investigation in the next funding period. Accordingly, this application outlines experiments designed to learn how desosamine, apiose, kijanose, and 2-deoxy-2- mercaptoglucose are biosynthesized. The specific objectives include (1) mechanistic studies of a radical-SAM enzyme (DesII) involved in the biosynthesis of desosamine, an essential component of many macrolide antibiotics, (2) a determination of the mechanism of the pyranose-to-furanose ring-contraction catalyzed by UDP-apiose synthase, (3) an investigation into the biosynthesis of the unusual nitrosugar moiety (kijanose) of kijanimicin, and (4) the elucidation of the mechanism of sulfur incorporation into the 2-deoxy-2-mercaptoglucose moiety of the antibiotic BE-7585A. The proposed experiments will not only delineate the biosynthesis of deoxy-, branched-chain, nitro-, and sulfur-containing sugars, but will also advance the field of mechanistic enzymology by enhancing our understanding of several important classes of enzymes. Our results should also be valuable to applied biomedical research, as new glycosylation tools (i.e., sugar biosynthetic pathways and enzymes) will be discovered for future secondary metabolite glycodiversification efforts.

Public Health Relevance

Outlined in this application are experiments designed to study the mechanism of the radical SAM enzyme, DesII, involved in the C4-deoxygenation step in desosamine biosynthesis, the mechanism of the ring contraction reaction catalyzed by UDP-apiose synthase (Axs1), the biosynthesis of an unusual nitrosugar, kijanose, and the mechanism of sulfur incorporation into the thiosugar moiety of BE-7585A. The results are expected to significantly advance the field of mechanistic enzymology and should also be valuable to applied biomedical research, as new sugar biosynthetic pathways and enzymes will be discovered for future secondary metabolite glycodiversification efforts.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM035906-28
Application #
8208088
Study Section
Macromolecular Structure and Function E Study Section (MSFE)
Program Officer
Gerratana, Barbara
Project Start
1986-01-01
Project End
2013-09-04
Budget Start
2012-01-01
Budget End
2013-09-04
Support Year
28
Fiscal Year
2012
Total Cost
$427,661
Indirect Cost
$138,569
Name
University of Texas Austin
Department
Pharmacology
Type
Schools of Pharmacy
DUNS #
170230239
City
Austin
State
TX
Country
United States
Zip Code
78712
Jackson, David R; Yu, Xia; Wang, Guojung et al. (2016) Insights into Complex Oxidation during BE-7585A Biosynthesis: Structural Determination and Analysis of the Polyketide Monooxygenase BexE. ACS Chem Biol 11:1137-47
Kim, Hak Joong; LeVieux, Jake; Yeh, Yu-Cheng et al. (2016) C3'-Deoxygenation of Paromamine Catalyzed by a Radical S-Adenosylmethionine Enzyme: Characterization of the Enzyme AprD4 and Its Reductase Partner AprD3. Angew Chem Int Ed Engl 55:3724-8
Bridwell-Rabb, Jennifer; Kang, Gyunghoon; Zhong, Aoshu et al. (2016) An HD domain phosphohydrolase active site tailored for oxetanocin-A biosynthesis. Proc Natl Acad Sci U S A 113:13750-13755
Ko, Yeonjin; Ruszczycky, Mark W; Choi, Sei-Hyun et al. (2015) Mechanistic studies of the radical S-adenosylmethionine enzyme DesII with TDP-D-fucose. Angew Chem Int Ed Engl 54:860-3
Lin, Geng-Min; Choi, Sei-Hyun; Ruszczycky, Mark W et al. (2015) Mechanistic Investigation of the Radical S-Adenosyl-L-methionine Enzyme DesII Using Fluorinated Analogues. J Am Chem Soc 137:4964-7
Hashimoto, Takuya; Hashimoto, Junko; Teruya, Kuniko et al. (2015) Biosynthesis of versipelostatin: identification of an enzyme-catalyzed [4+2]-cycloaddition required for macrocyclization of spirotetronate-containing polyketides. J Am Chem Soc 137:572-5
Ruszczycky, Mark W; Liu, Hung-Wen (2015) Mechanistic Enzymology of the Radical SAM Enzyme DesII. Isr J Chem 55:315-324
Fage, Christopher D; Isiorho, Eta A; Liu, Yungnan et al. (2015) The structure of SpnF, a standalone enzyme that catalyzes [4 + 2] cycloaddition. Nat Chem Biol 11:256-8
Kim, Hak Joong; Choi, Sei-hyun; Jeon, Byung-sun et al. (2014) Chemoenzymatic synthesis of spinosyn A. Angew Chem Int Ed Engl 53:13553-7
Isiorho, Eta A; Jeon, Byung-Sun; Kim, Nam Ho et al. (2014) Structural studies of the spinosyn forosaminyltransferase, SpnP. Biochemistry 53:4292-301

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