2 research outputs found
Cellular localization of D-lactate dehydrogenase and NADH oxidase from Archaeoglobus fulgidus
Members of the genus Archaeoglobus are
hyperthermophilic sulfate reducers with an optimal growth temperature
of 83 °C. Archaeoglobus fulgidus can utilize
simple compounds including D-lactate, L-lactate and pyruvate as the
sole substrate for carbon and electrons for dissimilatory sulfate
reduction. Previously we showed that this organism makes a D-lactate
dehydrogenase (Dld) that requires FAD and Zn2+ for
activity. To determine the cellular location and topology of Dld and
to identify proteins that interact with Dld, an antibody directed
against Dld was prepared. Immunocytochemical studies using gold
particle-coated secondary antibodies show that more than 85% of Dld is
associated with the membrane. A truncated form of Dld was detected in
immunoblots of whole cells treated with protease, showing that Dld is
an integral membrane protein and that a significant portion of Dld,
including part of the FAD-binding pocket, is outside the membrane
facing the S-layer. The gene encoding Dld is part of an operon that
includes noxA2, which encodes one of several NADH
oxidases in A. fulgidus. Previous studies have shown
that NoxA2 remains bound to Dld during purification. Thin sections of
A. fulgidus probed simultaneously with antibodies
against Dld and NoxA2 show that both proteins co-localized to the same
sites in the membrane. Although these data show a tight interaction
between NoxA2 and Dld, the role of NoxA2 in electron transport
reactions is unknown. Rather, NoxA2 may protect proteins involved in
electron transfer by reducing O2 to
H2O2 or H2O