Oligomeric Changes Regulate Flavin Transfer in Two-Component FMN Reductases Involved in Sulfur Metabolism

The FMN reductases (SsuE and MsuE of the alkanesulfonate
monooxygenase
systems) supply reduced flavin to their partner monooxygenases for
the desulfonation of alkanesulfonates. Flavin reductases that comprise
two-component systems must be able to regulate both flavin reduction
and transfer. One mechanism to control these distinct processes is
through changes in the oligomeric state of the enzymes. Despite their
similar overall structures, SsuE and MsuE showed clear differences
in their oligomeric states in the presence of substrates. The oligomeric
state of SsuE was converted from a tetramer to a dimer/tetramer equilibrium
in the presence of FMN or NADPH in analytical ultracentrifugation
studies. Conversely, MsuE shifted from a dimer to a single tetrameric
state with FMN, and the NADPH substrate did not induce a similar oligomeric
shift. There was a fast tetramer to dimer equilibrium shift occurring
at the dimer/dimer interface in H/D-X investigations with apo SsuE.
Formation of the SsuE/FMN complex slowed the tetramer/dimer conversion,
leading to a slower exchange along the dimer/dimer interface. The
oligomeric shift of the MsuE/FMN complex from a dimer to a distinct
tetramer showed a decrease in H/D-X in the region around the π-helices
at the dimer/dimer interface. Both SsuE and MsuE showed a comparable
and significant increase in the melting temperature with the addition
of FMN, indicating the conformers formed by each FMN-bound enzyme
had increased stability. A mechanism that supports the different structural
shifts is rationalized by the different roles these enzymes play in
providing reduced flavin to single or multiple monooxygenase enzymes.