1 research outputs found
Modulating the Molybdenum Coordination Sphere of <i>Escherichia coli</i> Trimethylamine <i>N</i>‑Oxide Reductase
The
well-studied enterobacterium <i>Escherichia coli</i> present
in the human gut can reduce trimethylamine <i>N</i>-oxide
(TMAO) to trimethylamine during anaerobic respiration. The
TMAO reductase TorA is a monomeric, bis-molybdopterin guanine dinucleotide
(bis-MGD) cofactor-containing enzyme that belongs to the dimethyl
sulfoxide reductase family of molybdoenzymes. We report on a system
for the <i>in vitro</i> reconstitution of TorA with molybdenum
cofactors (Moco) from different sources. Higher TMAO reductase activities
for TorA were obtained when using Moco sources containing a sulfido
ligand at the molybdenum atom. For the first time, we were able to
isolate functional bis-MGD from <i>Rhodobacter capsulatus</i> formate dehydrogenase (FDH), which remained intact in its isolated
state and after insertion into apo-TorA yielded a highly active enzyme.
Combined characterizations of the reconstituted TorA enzymes by electron
paramagnetic resonance spectroscopy and direct electrochemistry emphasize
that TorA activity can be modified by changes in the Mo coordination
sphere. The combination of these results together with studies of
amino acid exchanges at the active site led us to propose a novel
model for binding of the substrate to the molybdenum atom of TorA