Abstract
FocA is a predicted formate channel with a deduced mass of 31 kDa that catalyzes the bidirectional movement of formate across the cytoplasmic membrane of Escherichia coli and is the archetype of the formate-nitrite transporter (FNT) family. Overproduced FocA variants with either an N- or a C-terminal Strep-tag increased formate import into anaerobic E. coli cells as determined by the enhanced activity of a single-copy formate-dependent fdhF::lacZ fusion. Using anti-FocA antibodies, we could show that both FocA variants were integrated into the cytoplasmic membrane. Circular dichroism spectroscopy of purified FocA(Strep-N) revealed a high alpha-helical content of 56% consistent with the predicted six transmembrane helices present in the protein. Analysis of the oligomeric state by blue-native polyacrylamide gel electrophoresis revealed FocA to have an unexpected pentameric quaternary structure. This study reports the first isolation of an FNT family member.
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