Discovery and structural characterization of a thermostable bacterial monoamine oxidase

Monoamine oxidases (MAOs) are pivotal regulators of neurotransmitters
in mammals, while microbial MAOs have been shown to be valuable bio-
catalysts for enantioselective synthesis of pharmaceutical compounds or
precursors thereof. To extend the knowledge of how MAOs function at the
molecular level and in order to provide more biocatalytic tools, we set out
to identify and study a robust bacterial variant: a MAO from the thermo-
phile Thermoanaerobacterales bacterium (MAOTb). MAOTb is highly ther-
mostable with melting temperatures above 73 °C and is well expressed in
Escherichia coli. Substrate screening revealed that the oxidase is most effi-
cient with n-alkylamines with n-heptylamine being the best substrate.
Presteady-state kinetic analysis shows that reduced MAOTb rapidly reacts
with molecular oxygen, confirming that it is a bona fide oxidase. The crys-
tal structure of MAOTb was resolved at 1.5 A˚ and showed an exceptionally
high similarity with the two human MAOs, MAO A and MAO B. The
active site of MAOTb resembles mostly the architecture of human MAO A,
including the cysteinyl protein–FAD linkage. Yet, the bacterial MAO lacks
a C-terminal extension found in human MAOs, which explains why it is
expressed and purified as a soluble protein, while the mammalian counter-
parts are anchored to the membrane through an α-helix. MAOTb also dis-
plays a slightly different active site access tunnel, which may explain the
specificity toward long aliphatic amines. Being an easy-to-express, thermo-
stable enzyme, for which a high-resolution structure was elucidated, this
bacterial MAO may develop into a valuable biocatalyst for synthetic chem-
istry or biosensing.