title: Biosynthetic origin of hygromycin A.
abstract: Hygromycin A, an antibiotic produced by Streptomyces hygroscopicus, is an inhibitor of bacterial ribosomal peptidyl transferase. The antibiotic binds to the ribosome in a distinct but overlapping manner with other antibiotics and offers a different template for generation of new agents effective against multidrug-resistant pathogens. Reported herein are the results from a series of stable-isotope-incorporation studies demonstrating the biosynthetic origins of the three distinct structural moieties which comprise hygromycin A. Incorporation of [1-(13)C]mannose and intact incorporation of D-[1,2-(13)C(2)]glucose into the 6-deoxy-5-keto-D-arabino-hexofuranose moiety are consistent with a pathway in which mannose is converted to an activated L-fucose, via a 4-keto-6-deoxy-D-mannose intermediate, with a subsequent unusual mutation of the pyranose to the corresponding furanose. The aminocyclitol moiety was labeled by D-[1,2-(13)C(2)]glucose in a manner consistent with formation of myo-inositol and a subsequent unprecedented oxidation and transamination of the C-2 hydroxyl group to generate neo-inosamine-2. Incorporation of [carboxy-(13)C]-4-hydroxybenzoic acid and intact incorporation of [2,3-(13)C(2)]propionate are consistent with a polyketide synthase-type decarboxylation condensation to generate the 3,4-dihydroxy-alpha-methylcinnamic acid moiety of hygromycin A. No labeling of hygromycin A was observed when [3-(13)C]tyrosine, [3-(13)C]phenylalanine, or [carboxy-(13)C]benzoic acid was used, suggesting that the 4-hydroxybenzoic acid is derived directly from chorismic acid. Consistent with this hypothesis was the observation that hygromycin A titers could be reduced by addition of N-(phosphonomethyl)-glycine (an inhibitor of chorismic acid biosynthesis) and restored by coaddition of 4-hydroxybenzoic acid. The convergent biosynthetic pathway established for hygromycin A offers significant versatility for applying the techniques of combinatorial and directed biosynthesis to production of new antibiotics which target the ribosomal peptidyl transferase activity.
authors.affiliation: Departments of Medicinal Chemistry,Virginia Commonwealth University, Richmond, Virginia 23219, USA.
authors.forename: El-Sayed E
authors.initials: el-SE
authors.lastname: Habib
authors.forename: J Neel
authors.initials: JN
authors.lastname: Scarsdale
authors.forename: Kevin A
authors.initials: KA
authors.lastname: Reynolds
mesh_terms: D000900:Anti-Bacterial Agents / Q000096:biosynthesis* / Q000737:chemistry; D002934:Cinnamates / Q000737:chemistry; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005285:Fermentation; D005998:Glycine / Q000031:analogs & derivatives* / Q000494:pharmacology; D006921:Hygromycin B / Q000031:analogs & derivatives* / Q000096:biosynthesis* / Q000737:chemistry; D013302:Streptomyces / Q000378:metabolism*; D000097797:Glyphosate
chemical_list: D000900:Anti-Bacterial Agents; D002934:Cinnamates; D004791:Enzyme Inhibitors; D005998:Glycine; D006921:Hygromycin B; D000097797:Glyphosate; C029010:cinnamic acid; C026273:hygromycin A