Stability and folding of the ferredoxin from the hyperthermophilic archaeon Acidianus ambivalens.
The ferredoxin from the thermophilic archaeon Acidianus ambivalens is a small monomeric protein containing two iron-sulfur centres, one [3Fe-4S](1+/0) and one [4Fe-4S](2+/1+). It is an intrinsically hyperstable protein, being expressed at the organism's extreme optimal growth temperature: 80 degrees C. Using spectroscopic methods we have investigated the unfolding reaction of the Acidianus ambivalens ferredoxin. No unfolding of the oxidised ferredoxin was observed at pH 7.0, even in the presence of 8 M GuHCl. Upon increasing the pH to 10.0, the unfolding transition showed a midpoint at 6.3 M GuHCl and an unfolding-free energy of 70 kJ mol(-1) in buffer (pH 10) was estimated. Kinetic-unfolding experiments showed that the polypeptide unfolding correlated with rearrangement of the iron-sulfur centres to new ones which had strong absorption maxima at 520 and 610 nm. These new, possibly linear three-iron, clusters were coordinated to the unfolded protein but degraded slowly. From thermal experiments in the presence of GuHCl we estimated the melting temperature for the Acidianus ambivalens ferredoxin in buffer (at pH 7) to be 122 degrees C. Possible structural properties that contribute to the large thermal stability of the Acidianus ambivalens ferredoxin are discussed using a three-dimensional protein model.
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Chemistry Department, Tulane University, New Orleans, LA 70118, USA. pernilla@mailhost.tcs.tulane.edu
Gomes
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D001105:Archaea / Q000737:chemistry*; D019843:Archaeal Proteins / Q000737:chemistry; D002942:Circular Dichroism; D003116:Color; D004227:Dithionite / Q000494:pharmacology; D004578:Electron Spin Resonance Spectroscopy; D005288:Ferredoxins / Q000737:chemistry*; D019791:Guanidine / Q000494:pharmacology; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007501:Iron / Q000737:chemistry; D007506:Iron-Sulfur Proteins / Q000737:chemistry; D007700:Kinetics; D008958:Models, Molecular; D015394:Molecular Structure; D010084:Oxidation-Reduction; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D013053:Spectrophotometry; D013056:Spectrophotometry, Ultraviolet; D013455:Sulfur / Q000737:chemistry; D013696:Temperature
D019843:Archaeal Proteins; D005288:Ferredoxins; D007506:Iron-Sulfur Proteins; D004227:Dithionite; D013455:Sulfur; D007501:Iron; D019791:Guanidine