SEQUENCE, ASSEMBLY AND EVOLUTION OF A PRIMORDIAL FERREDOXIN FROM THERMOTOGA-MARITIMA

A gene coding for the ferredoxin of the primordial, strictly anaerobic and hyperthermophilic bacterium Thermotoga maritima was cloned, seque nced and expressed in Escherichia coli. The ferredoxin gene encodes a polypeptide of 60 amino acids that incorporates a single 4Fe-4S cluste r. T.maritima ferredoxin expressed in E.coli is a heat-stable, monomer ic protein, the spectroscopic properties of which show that its 4Fe-4S cluster is correctly assembled within the mesophilic host, and that i t remains stable during purification under aerobic conditions. Removal of the iron-sulfur cluster results in an apo-ferredoxin that has no d etectable secondary structure. This observation indicates that in vivo formation of the ferredoxin structure is coupled to the insertion of the iron-sulfur cluster into the polypeptide chain. Sequence compariso n of T.maritima ferredoxin with other 4Fe-4S ferredoxins revealed high sequence identities (75% and 50% respectively) to the ferredoxins fro m the hyperthermophilic members of the Archaea, Thermococcus litoralis and Pyrococcus furiosus. The high sequence similarity supports a clos e relationship between these extreme thermophilic organisms from diffe rent phylogenetic domains and suggests that ferredoxins with a single 4Fe-4S cluster are the primordial representatives of the whole protein family. This observation suggests a new model for the evolution of fe rredoxins.