ISOLATION AND CHARACTERIZATION OF METHANOPHENAZINE AND FUNCTION OF PHENAZINES IN MEMBRANE-BOUND ELECTRON-TRANSPORT OF METHANOSARCINA-MAZEI GOL

A hydrophobic, redox-active component with a molecular mass of 538 Da was isolated from lyophilized membranes of Methanosarcina mazei Gol by extraction with isooctane, After purification on a high-performance l iquid chromatography column, the chemical structure was analyzed by ma ss spectroscopy and nuclear magnetic resonance studies. The component was called methanophenazine and represents a 2-hydrophenazine derivati ve which is connected via an ether bridge to a polyisoprenoid side cha in. Since methanophenazine was almost insoluble in aqueous buffers, wa ter-soluble phenazine derivatives were tested for their ability to int eract with membrane-bound enzymes involved in electron transport and e nergy conservation. The purified F420H2 dehydrogenase from M. mazei Go 1 showed highest activity with 2-hydroxyphenazine and 2-bromophenazine as electron acceptors when F420H2 was added. Phenazine-1-carboxylic a cid and phenazine proved to be less effective. The K-m values for 2-hy droxyphenazine and phenazine were 35 and 250 mu M, respectively. 2-Hyd roxyphenazine was also reduced by molecular hydrogen catalyzed by an F -420-nonreactive hydrogenase which is present in washed membrane prepa rations. Furthermore, the membrane-bound heterodisulfide reductase was able to use reduced 2-hydroxyphenazine as an electron donor for the r eduction of CoB-S-S-CoM. Considering all these results, it is reasonab le to assume that methanophenazine plays an important role in vivo in membrane-bound electron transport of M. mazei Go1.