2020
DOI: 10.1002/ange.202005554
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In Vitro Reconstitution Reveals a Central Role for the N‐Oxygenase PvfB in (Dihydro)pyrazine‐N‐oxide and Valdiazen Biosynthesis

Abstract: The Pseudomonas virulence factor (pvf) operon is essential for the biosynthesis of two very different natural product scaffolds: the (dihydro)pyrazine-N-oxides and the diazeniumdiolate, valdiazen. PvfB is a member of the nonheme diiron N-oxygenase enzyme family that commonly convert anilines to their nitroaromatic counterparts. In contrast, we show that PvfB catalyzes N-oxygenation of the aamine of valine, first to the hydroxylamine and then the nitroso, while linked to the carrier protein of PvfC. PvfB modifi… Show more

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Cited by 5 publications
(5 citation statements)
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“…160,161 Indeed, pvfB and pvfC were demonstrated to be necessary and sufficient for the production of 207-209 in vivo. Recently, the N-hydroxylation mechanism was shown by protein NMR analysis to operate in both fragin and diisopropylpyrazine pyrazine N-oxides biosynthesis as follows: the putative diiron N-oxygenase PvfB/HamC, catalyzes the 2-electron oxidation of valine tethered PCP domain of PvfC/HamD to form a hydroxylamine intermediate 162 (Fig. 32D).…”
Section: Biosynthesis Of N-nitroso Compoundsmentioning
confidence: 99%
“…160,161 Indeed, pvfB and pvfC were demonstrated to be necessary and sufficient for the production of 207-209 in vivo. Recently, the N-hydroxylation mechanism was shown by protein NMR analysis to operate in both fragin and diisopropylpyrazine pyrazine N-oxides biosynthesis as follows: the putative diiron N-oxygenase PvfB/HamC, catalyzes the 2-electron oxidation of valine tethered PCP domain of PvfC/HamD to form a hydroxylamine intermediate 162 (Fig. 32D).…”
Section: Biosynthesis Of N-nitroso Compoundsmentioning
confidence: 99%
“…26 HamC, which shares homology with diiron N -oxygenase AurF, 27 was found to be responsible for the in vitro nitrogen oxidation of valine, while the substrate was bound to the NRPS complex. 11,23,28 The cupin domain of HamB was suggested nonessential for the production of valdiazen, 11 and that HamA (PvfA) and HamE (PvfD) were nonessential for the production of hydroxylamine intermediates towards valdiazen and pyrazine N -oxide derivatives. 22 From the second operon, HamG was predicted to be an aminotransferase, and HamF a starter condensation domain, 11 and both of them are indispensable for the production of fragin.…”
Section: Resultsmentioning
confidence: 99%
“…fragin and (±)-valdiazen, based on previous findings from our group and others. 11,[13][14][15][16][19][20][21][22][23][24][25][26] The ham cluster consists of seven genes distributed in two oppositely oriented operons. 11,26 As demonstrated by isotopically labelled feeding experiments, the biosynthesis starts with loading L-valine onto the HamD nonribosomal peptide synthetase (NRPS) complex.…”
Section: Introductionmentioning
confidence: 99%
“…However, general efficient synthetic routes to access diazeniumdiolate analogs were missing in the literature. While the biosynthesis of some natural diazeniumdiolates has been investigated, 19,20,47,5052,54–60 only a limited number of synthetic derivatives have been described. 1820,61 In the case of leudiazen, the previous synthesis in 4 steps from DL- leucinol yielded only a 15% overall yield (Scheme 1A), 12 being far from efficient.…”
Section: Resultsmentioning
confidence: 99%