Part:BBa_K535006
PFOR -> D. africanus Pyruvate-Ferredoxin OxidoReductase
Pyruvate-ferredoxin oxidoreductases (PFORs) are a widespread group of proteins from several anaerobic or microaerobic species of microorganisms, with a variety of functions in energy metabolism, and CO2 fixation.
Though they generally catalyze the crucial energy-yielding reaction of pyruvate oxidation and serve as electron carriers, sometimes they differ in distinct species in their acceptor/donor specificity, substrate specificity, sensitivity to O2, and kinetics of CO2 production and consumption. They are complex iron-sulfur proteins containing thiamine pyrophosphate (TPP) and one or two iron-sulfur [4Fe-4S] clusters. They contain neither flavin nor lipoate as prosthetic groups.
This sequence corresponds to the PFOR from Desulfovibrio africanus which is a Gram negative sulfate-reducing bacterium, mostly inhabiting anaerobic environments. PFOR in this organism has been reported to be located in the cytoplasm, like the reductases of the sulfate reduction pathway.
The specific activity of D. africanus PFOR is exceptionally high compared with other mesophile PFORs reported.
D. africanus' PFOR is a 256±10 kDa homodimer which contains two thiamine pyrophosphate (TPP). Each subunit has three [4Fe-4S] 2+/1+ centers, which is higher than other PFORs which have at most two [4Fe-4S] clusters per subunit. The midpoint potentials of the three centers are -390 mV, -515 mV and -540 mV.
The three [4Fe-4S] clusters are close enough in the PFOR subunit to interact magnetically, namely less than about 1.5 nm apart.
In Desulfovibrio species the enzyme is involved in the oxidation of pyruvate and provides the sulfate reducing pathway with low potential electrons. Generally, ferredoxin and flavodoxin serve as electron acceptors for the enzyme. Ferredoxins are the most effective acceptor for the D. africanus' enzyme.
In order to work, the enzyme should be activated by dithioerythritol. Neither the nature, nor the arrangement of [4Fe-4S] centers are affected by the activation process, nevertheless there is a change in the protein's conformation during activation which makes the enzyme very sensitive to oxygen in contrast to the enzyme before activation. This is believed to happen given that iron-sulfur clusters are now exposed to oxidative damage.
In our project we intend to create a pathway with heterologous expressed PFOR-ferredoxin-hydrogenase in order to couple the hydrogen production to the glycolisis process.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NotI site found at 1833
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 146
Illegal NgoMIV site found at 359
Illegal NgoMIV site found at 368
Illegal NgoMIV site found at 395
Illegal NgoMIV site found at 1117
Illegal NgoMIV site found at 1244
Illegal NgoMIV site found at 1279 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 2131
Illegal BsaI.rc site found at 1007
Illegal BsaI.rc site found at 2048
Illegal BsaI.rc site found at 2615
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