4.1 Cytoplasmic Membrane
Electron transfer at the cytoplasmic membrane involves the linkage of dehydrogenases responsible for oxidation of carbon sources in the cytoplasm (e.g. formate
dehydrogenase), through a lipid soluble quinone pool, to electron transfer proteins
(e.g. cytochromes) bound to the cytoplasmic membrane [84]. This mechanism
generates a proton-electrochemical gradient that is used to produce ATP via the
ATP synthase [85]. In parallel, the electron flow through the quinone pool towards
extracellular electron acceptors appears to serve also as a pathway to discharge
electrons without coupling to the generation of transmembrane electrochemical
potential [86].
S. oneidensis MR-1 is known to produce three quinones (menaquinone,
methylmenaquinone and ubiquinone) [87]. The deletion of menD and menB genes
involved in the biosynthesis of menaquinone produced a phenotype incapable of
iron respiration, revealing that menaquinone but not ubiquinone plays a role in
metal respiration [88, 89]. This is in line with the difference in the reduction
potentials of these two quinones with the ubiquinone potential more aligned to
participate in aerobic respiratory chains.
4.1.1 CymA
Presently, it is well established that the linkage between the membrane quinone
pool and EET chain is provided by a tetraheme c-type cytochrome called CymA
that is attached to the periplasmic surface of the cytoplasmic membrane by a ahelical anchor [88, 90–93]. CymA from Shewanella has 21 kDa and is a member of
the NapC/NirT protein family. It is able to bind quinol (Kd = 0.1–1 lM) [94],
functioning as a quinol oxidase [88, 92, 95]. Deletion of the cymA gene severely
hindered the reduction of a variety of substrates including Fe(III)/Mn(IV) oxides,
fumarate, nitrate, nitrate, and DMSO [82, 90, 91]. This supported the proposal that
CymA is one of the major hubs for electron transfer to the periplasm, being
essential for EET. CymA’s ability to interact with multiple periplasmic cytochrome
partners has been amply demonstrated and explored [92, 96, 97]. Also, overexpression of this gene is enough to enhance electricity generation by S. oneidensis
MR-1 in an MFC [98]. CymA’s role in EET was further confirmed by cloning the
gene in E. coli and observing that the heterologous expression of CymA is enough
to make this bacterium capable of EET [99].
Although no structural characterization is presently available for CymA, it
contains three low spin hemes with bis–histidine axial ligation and one high-spin
heme with a histidine–water axial ligation [88, 100]. This high-spin heme forms an
intrinsic part of the quinol oxidation site. Also, site-directed mutagenesis experiments revealed that the amino acid Lysine-91 is essential for quinol interaction with
CymA from Shewanella sp. strain ANA-3 [94]. Redox properties were determined
for CymA from S. oneidensis MR-1, with macroscopic midpoint potentials at pH
7.0 of approximately −110, −190 and −265 mV for the three low-spin hemes and
−240 mV for the high-spin heme [88]. These potentials are below that of the
menaquinol/menaquinone couple (Eº’ % −80 mV), and thus electron transfer only
224
B. M. Fonseca et al.
Electron transfer at the cytoplasmic membrane involves the linkage of dehydrogenases responsible for oxidation of carbon sources in the cytoplasm (e.g. formate
dehydrogenase), through a lipid soluble quinone pool, to electron transfer proteins
(e.g. cytochromes) bound to the cytoplasmic membrane [84]. This mechanism
generates a proton-electrochemical gradient that is used to produce ATP via the
ATP synthase [85]. In parallel, the electron flow through the quinone pool towards
extracellular electron acceptors appears to serve also as a pathway to discharge
electrons without coupling to the generation of transmembrane electrochemical
potential [86].
S. oneidensis MR-1 is known to produce three quinones (menaquinone,
methylmenaquinone and ubiquinone) [87]. The deletion of menD and menB genes
involved in the biosynthesis of menaquinone produced a phenotype incapable of
iron respiration, revealing that menaquinone but not ubiquinone plays a role in
metal respiration [88, 89]. This is in line with the difference in the reduction
potentials of these two quinones with the ubiquinone potential more aligned to
participate in aerobic respiratory chains.
4.1.1 CymA
Presently, it is well established that the linkage between the membrane quinone
pool and EET chain is provided by a tetraheme c-type cytochrome called CymA
that is attached to the periplasmic surface of the cytoplasmic membrane by a ahelical anchor [88, 90–93]. CymA from Shewanella has 21 kDa and is a member of
the NapC/NirT protein family. It is able to bind quinol (Kd = 0.1–1 lM) [94],
functioning as a quinol oxidase [88, 92, 95]. Deletion of the cymA gene severely
hindered the reduction of a variety of substrates including Fe(III)/Mn(IV) oxides,
fumarate, nitrate, nitrate, and DMSO [82, 90, 91]. This supported the proposal that
CymA is one of the major hubs for electron transfer to the periplasm, being
essential for EET. CymA’s ability to interact with multiple periplasmic cytochrome
partners has been amply demonstrated and explored [92, 96, 97]. Also, overexpression of this gene is enough to enhance electricity generation by S. oneidensis
MR-1 in an MFC [98]. CymA’s role in EET was further confirmed by cloning the
gene in E. coli and observing that the heterologous expression of CymA is enough
to make this bacterium capable of EET [99].
Although no structural characterization is presently available for CymA, it
contains three low spin hemes with bis–histidine axial ligation and one high-spin
heme with a histidine–water axial ligation [88, 100]. This high-spin heme forms an
intrinsic part of the quinol oxidation site. Also, site-directed mutagenesis experiments revealed that the amino acid Lysine-91 is essential for quinol interaction with
CymA from Shewanella sp. strain ANA-3 [94]. Redox properties were determined
for CymA from S. oneidensis MR-1, with macroscopic midpoint potentials at pH
7.0 of approximately −110, −190 and −265 mV for the three low-spin hemes and
−240 mV for the high-spin heme [88]. These potentials are below that of the
menaquinol/menaquinone couple (Eº’ % −80 mV), and thus electron transfer only
224
B. M. Fonseca et al.