Nos proteins specifically encoded in clade II NGCs are NosB, -G, -C1, -C2 and -
H (Table 2). The nosB gene is typically surrounded by nosZ and nosD (Fig. 2) [18,
37, 70]. NosB proteins are predicted to traverse the membrane four (in some cases
six) times [45]. In W. succinogenes, characterization of a non-polar nosB deletion
mutant demonstrated that NosB was essential for N 2 O respiration [45]. Apart from
this, functional studies on NosB are not available.
NosG and NosH are likely to form a membrane-bound complex involved in
(transmembrane) electron transport (Fig. 3b). NosG is a periplasmic iron-sulphur
protein that is exported by the Tat pathway and predicted to bind four [4Fe–4S]
centres, whereas NosH is a membrane-bound protein predicted to form four
transmembrane segments [45, 58]. Similar to NosR, NosH contains a C-terminal
cytoplasmic domain that putatively binds two [4Fe–4S] centres and two conserved
CX 3 CP motifs at the cytoplasmic side of the membrane. NosG and NosH are
homologues of NapG and NapH, two essential components of the periplasmic
nitrate reductase system (Nap) in various Proteobacteria [97–100]. Previously,
NosGH- and NapGH-type complexes have been postulated to be involved in
electron transport from the quinone/quinol pool to a terminal reductase of anaerobic
respiration (in this case cNosZ or the nitrate reductase NapA) [16, 58, 97]. However, direct biochemical evidence for quinol reactivity is lacking. NosC1 and
NosC2 are monohaem cytochromes c located either in the periplasm or attached to
the membrane via an N-terminal helix. These two cytochromes c might function as
redox partner proteins of NosG. Non-polar W. succinogenes single-gene deletion
mutants deficient in NosG, NosC1, NosC2 or NosH did not grow by N 2 O respiration, and this phenotype was successfully complemented for each mutant upon
genomic incorporation of the respective gene [45]. Thus, in principle, the NosG, -
C1, -C2, -H and -B proteins could be functionally and/or structurally involved in
electron transport from the QcrABC complex (which has been also shown to be
essential for N 2 O respiration in W. succinogenes) to cNosZ forming an elongated
electron transport chain when compared to that of clade I organisms [45]. However,
the experimental evidence is also in line with an alternative scenario, in which
NosH, -G, -C1 and -C2 functionally replace NosR (Fig. 3b). According to this
model, the absence of any of the NosH, -G, -C1 and -C2 proteins could be deleterious for N 2 O respiration as the electron transport route required for reductive
cNosZ activation and/or NosDL-dependent cNosZ biogenesis would be abolished.
Lack of NosB could lead to the same phenotype assuming that this protein provides
a membrane-bound platform for the correct assembly of the Nos system. Finally,
the cytochrome c domain of cNosZ could mediate the NosGH-initiated electron
transport route. This view is supported by the fact that the cytochrome c domain is
connected to NosZ by a surprisingly large linker sequence that possibly makes this
domain very flexible arguing for a transient interaction with the active site of NosZ
[58, 59].
Notably, various Nos fusion proteins (NosD-F, NosB-C2, NosB-L) are predicted
from (meta)genomic clade II NGC data (Table 1). In addition, NosL-D-type fusion
proteins have been described, which, however, are not encoded by genes organized
in NGCs [70]. The predicted multidomain proteins support the model shown in
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H (Table 2). The nosB gene is typically surrounded by nosZ and nosD (Fig. 2) [18,
37, 70]. NosB proteins are predicted to traverse the membrane four (in some cases
six) times [45]. In W. succinogenes, characterization of a non-polar nosB deletion
mutant demonstrated that NosB was essential for N 2 O respiration [45]. Apart from
this, functional studies on NosB are not available.
NosG and NosH are likely to form a membrane-bound complex involved in
(transmembrane) electron transport (Fig. 3b). NosG is a periplasmic iron-sulphur
protein that is exported by the Tat pathway and predicted to bind four [4Fe–4S]
centres, whereas NosH is a membrane-bound protein predicted to form four
transmembrane segments [45, 58]. Similar to NosR, NosH contains a C-terminal
cytoplasmic domain that putatively binds two [4Fe–4S] centres and two conserved
CX 3 CP motifs at the cytoplasmic side of the membrane. NosG and NosH are
homologues of NapG and NapH, two essential components of the periplasmic
nitrate reductase system (Nap) in various Proteobacteria [97–100]. Previously,
NosGH- and NapGH-type complexes have been postulated to be involved in
electron transport from the quinone/quinol pool to a terminal reductase of anaerobic
respiration (in this case cNosZ or the nitrate reductase NapA) [16, 58, 97]. However, direct biochemical evidence for quinol reactivity is lacking. NosC1 and
NosC2 are monohaem cytochromes c located either in the periplasm or attached to
the membrane via an N-terminal helix. These two cytochromes c might function as
redox partner proteins of NosG. Non-polar W. succinogenes single-gene deletion
mutants deficient in NosG, NosC1, NosC2 or NosH did not grow by N 2 O respiration, and this phenotype was successfully complemented for each mutant upon
genomic incorporation of the respective gene [45]. Thus, in principle, the NosG, -
C1, -C2, -H and -B proteins could be functionally and/or structurally involved in
electron transport from the QcrABC complex (which has been also shown to be
essential for N 2 O respiration in W. succinogenes) to cNosZ forming an elongated
electron transport chain when compared to that of clade I organisms [45]. However,
the experimental evidence is also in line with an alternative scenario, in which
NosH, -G, -C1 and -C2 functionally replace NosR (Fig. 3b). According to this
model, the absence of any of the NosH, -G, -C1 and -C2 proteins could be deleterious for N 2 O respiration as the electron transport route required for reductive
cNosZ activation and/or NosDL-dependent cNosZ biogenesis would be abolished.
Lack of NosB could lead to the same phenotype assuming that this protein provides
a membrane-bound platform for the correct assembly of the Nos system. Finally,
the cytochrome c domain of cNosZ could mediate the NosGH-initiated electron
transport route. This view is supported by the fact that the cytochrome c domain is
connected to NosZ by a surprisingly large linker sequence that possibly makes this
domain very flexible arguing for a transient interaction with the active site of NosZ
[58, 59].
Notably, various Nos fusion proteins (NosD-F, NosB-C2, NosB-L) are predicted
from (meta)genomic clade II NGC data (Table 1). In addition, NosL-D-type fusion
proteins have been described, which, however, are not encoded by genes organized
in NGCs [70]. The predicted multidomain proteins support the model shown in
Mitigation of Laughing Gas Emissions …
199