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Title
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PhzA/B catalyzes the formation of the tricycle in phenazine biosynthesis.
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Authors
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E.G.Ahuja,
P.Janning,
M.Mentel,
A.Graebsch,
R.Breinbauer,
W.Hiller,
B.Costisella,
L.S.Thomashow,
D.V.Mavrodi,
W.Blankenfeldt.
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Ref.
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J Am Chem Soc, 2008,
130,
17053-17061.
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PubMed id
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Abstract
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Phenazines are redox-active bacterial secondary metabolites that participate in
important biological processes such as the generation of toxic reactive oxygen
species and the reduction of environmental iron. Their biosynthesis from
chorismic acid depends on enzymes encoded by the phz operon, but many details of
the pathway remain unclear. It previously was shown that phenazine biosynthesis
involves the symmetrical head-to-tail double condensation of two identical
amino-cyclohexenone molecules to a tricyclic phenazine precursor. While this key
step can proceed spontaneously in vitro, we show here that it is catalyzed by
PhzA/B, a small dimeric protein of the Delta(5)-3-ketosteroid isomerase/nuclear
transport factor 2 family, and we reason that this catalysis is required in
vivo. Crystal structures in complex with analogues of the substrate and product
suggest that PhzA/B accelerates double imine formation by orienting two
substrate molecules and by neutralizing the negative charge of tetrahedral
intermediates through protonation. HPLC-coupled NMR reveals that the
condensation product rearranges further, which is probably important to prevent
back-hydrolysis, and may also be catalyzed within the active site of PhzA/B. The
rearranged tricyclic product subsequently undergoes oxidative decarboxylation in
a metal-independent reaction involving molecular oxygen. This conversion does
not seem to require enzymatic catalysis, explaining why phenazine-1-carboxylic
acid is a major product even in strains that use phenazine-1,6-dicarboxylic acid
as a precursor of strain-specific phenazine derivatives.
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