Enzymic synthesis of the 4Fe-4S clusters of Clostridium pasteurianum ferredoxin.
Ex novo enzymic synthesis of the two 4Fe-4S clusters of Clostridium pasteurianum ferredoxin has been achieved by incubation of the apoprotein with catalytic amounts of the sulfurtransferase rhodanese in the presence of thiosulfate, DL-dihydrolipoate and ferric ammonium citrate. This enzymic reconstitution procedure was compared to a chemical one, in which the enzyme was replaced by sodium sulfide. A further comparison was made with the results previously obtained in the enzymic synthesis of the 2Fe-2S cluster of spinach ferredoxin, allowing the following conclusions to be drawn. The nature of the cluster to be inserted into the reconstituted iron-sulfur protein is determined by the apoprotein itself. The refolding of the structure of the iron-sulfur proteins around the newly inserted cluster is the rate-limiting step in both chemical and enzymic reconstitution. Rhodanese appears to play a role in the recovery of the native architecture of the reconstituted iron-sulfur protein(s). The extension to the 4Fe-4S centers of the rhodanese-based biosynthetic system allows this enzymic route to be proposed as a general way to the in vivo synthesis of iron-sulfur structures.
Bonomi
F
F
Pagani
S
S
Kurtz
D M
DM
D001059:Apoproteins / Q000378:metabolism; D002384:Catalysis; D002942:Circular Dichroism; D003013:Clostridium / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy; D005288:Ferredoxins / Q000096:biosynthesis*; D013466:Sulfurtransferases / Q000378:metabolism*; D013884:Thiosulfate Sulfurtransferase / Q000378:metabolism*
D001059:Apoproteins; D005288:Ferredoxins; D013466:Sulfurtransferases; D013884:Thiosulfate Sulfurtransferase