Isolation, functional characterization and crystallization of Aq_1259, an outer membrane protein with porin features, from Aquifex aeolicus

The "hypothetical protein" Aq_1259 was identified by mass spectrometry and purified from native membranes of Aquifex aeolicus. It is a 49.4kDa protein, highly homologous (>52% identity) to several conserved hypothetical proteins from other bacteria. However, none of these proteins has been characterized using biochemical or electrophysiological techniques. Based on the sequence and circular dichroism spectroscopy, the structure of Aq_1259 is predicted to be a β-barrel with 16 β-strands. The strands with loops and turns are distributed evenly through the entire sequence. The function of Aq_1259 was analyzed after incorporation into a lipid bilayer. Electrophysiological measurements revealed a pore that has a basic stationary conductance of 0.48 ± 0.038nS in a buffer with 0.5M NaH₂PO₄ at pH 6.5 and 0.2 ± 0.015nS in a buffer with 0.5M NaCl at pH 6.5. Superimposed on this is a fluctuating conductance of similar amplitude. Aq_1259 could be crystallized. The crystals diffract to a resolution of 3.4Å and belong to space group I222 with cell dimensions of a=138.3Å, b=144.6Å, c=151.8Å.