title: Olefin isomerization regiochemistries during tandem action of BacA and BacB on prephenate in bacilysin biosynthesis.
abstract: BacA and BacB, the first two enzymes of the bacilysin pathway, convert prephenate to an exocylic regioisomer of dihydrohydroxyphenylpyruvate (ex-H(2)HPP) on the way to the epoxycyclohexanone warhead in the dipeptide antibiotic, bacilysin. BacA decarboxylates prephenate without aromatization, converting the 1,4-diene in prephenate to the endocyclic 1,3-diene in Δ(4),Δ(8)-dihydrohydroxyphenylpyruvate (en-H(2)HPP). BacB then performs an allylic isomerization to bring the diene into conjugation with the 2-ketone in the product Δ(3),Δ(5)-dihydrohydroxyphenylpyruvate (ex-H(2)HPP). To prove that BacA acts regiospecifically on one of the two prochiral olefins in prephenate, we generated 1,5,8-[(13)C]-chorismate from bacterial fermentation of 5-[(13)C]-glucose and in turn produced 2,4,6-[(13)C]-prephenate via chorismate mutase. Tandem action of BacA and BacB gave 2,4,8-[(13)C]-7R-ex-H(2)HPP, showing that BacA isomerizes only the pro-R double bond in prephenate. Nonenzymatic isomerization of the BacA product into conjugation gives only the Δ(3)E-geometric isomer of Δ(3),Δ(5)-ex-H(2)HPP. On the other hand, acceleration of the allylic isomerization by BacB gives a mixture of the E- and Z-geometric isomers of the 7R- product, indicating some rerouting of the flux, likely through dienolate geometric isomers.
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, United States.
authors.forename: Jared B
authors.initials: JB
authors.lastname: Parker
authors.forename: Christopher T
authors.initials: CT
authors.lastname: Walsh
mesh_terms: D000475:Alkenes / Q000737:chemistry* / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D002826:Chorismate Mutase / Q000737:chemistry / Q000378:metabolism; D003509:Cyclohexanecarboxylic Acids / Q000737:chemistry / Q000378:metabolism; D053138:Cyclohexenes / Q000737:chemistry / Q000378:metabolism; D004151:Dipeptides / Q000096:biosynthesis / Q000737:chemistry; D007536:Isomerism
chemical_list: D000475:Alkenes; D001426:Bacterial Proteins; D003509:Cyclohexanecarboxylic Acids; D053138:Cyclohexenes; D004151:Dipeptides; C006674:bacilysin; D002826:Chorismate Mutase; C005550:prephenic acid