Small structural changes account for the high thermostability of 1[4Fe–4S] ferredoxin from the hyperthermophilic bacterium Thermotoga maritima

Abstract

A comparison of the three-dimensional structure of FdTm with that of ferredoxins from mesophilic organisms suggests that the very high thermostability of FdTm is unexpectedly achieved without large changes of the overall protein structure. Instead, an increased number of potentially stabilizing features is observed in FdTm, compared with mesophilic ferredoxins. These include stabilization of alpha helices, replacement of residues in strained conformation by glycines, strong docking of the N-terminal methionine and an overall increase in the number of hydrogen bonds. Most of these features stabilize several secondary structure elements and improve the overall rigidity of the polypeptide backbone. The decreased flexibility will certainly play a relevant role in shielding the iron-sulfur cluster against physiologically high temperatures and further improve the functional integrity of FdTm.

Physics and evolution of thermophilic adaptation

“…Here, all stabilizing structural factors act in concert, pointing to enhanced compactness as the most probable original cause for higher stability. In the second case, on the contrary, we found a strong sequence bias that can explain dominating role of some of the stabilizing interactions, e.g., electrostatics (19,20), but not of others. The high level of sequence variation compared with mesophilic orthologs and the significant bias toward charged residues in their sequences point to a key role of sequence selection in adaptation of T. maritima proteins (1TMY and 1VJW) to extreme conditions of the environment, in contrast to other (hyper)thermophilic proteins (1CAA, 1IQZ, 2CJN, and 2PRD) where structural bias is more pronounced.…”

“…3 and Table 5), in turn, uncovers another possible mechanism of thermostability in T. maritima's proteins. The stability of these proteins under extremely high temperatures is apparently provided by significant modifications of their sequences toward enrichment by charged residues (19,20), which can be an effective sequence-based method of adaptation to extreme specific conditions. van der Waals interactions (29), number of H-bonds (12,30), and number of residues involved into elements of secondary structure in groups of proteins under consideration.…”

“…The increase of the number of charged residues in hyperthermophilic proteomes appears to be much greater than would have been necessary for stability purposes alone. Indeed, enhanced stability can be achieved by the addition of only a few ion pairs (4,19,20). This points to the possibility of alternative reasons (unrelated to protein stabilization) for this specific compositional bias toward charged residues, which should be thoroughly explored.…”

“…1a) and structural analysis (Table 1), it points to compactness as a key factor in the structure-based strategy of thermophilic adaptation in archaea. At the same time, lower packing density in proteins from A. aeolicus and T. maritima and a higher (compared with mesophilic E. coli, as well as with hyperthermophilic P. furiosus) content of charged residues suggest that these organisms follow mostly sequence-based stabilization, possibly, with a key role of ionic interactions (19,20).…”

The D13C variant of Bacillus schlegelii 7Fe ferredoxin is an 8Fe ferredoxin as revealed by 1H‐NMR spectroscopy

“…The latter resonance is characterized by its NOE to a resonance at 4.24 ppm, presumably Asp 23 Ha. This assignment is supported by the fact that the structurally characterized cubane ferredoxins share a turn involving the fourth cysteine of the cluster binding sequence (C(IV)PXX) as a highly conserved structural element [16][17][18][19][20][21][22][23]. Owing to this turn, a strong NOE connectivity is expected between CysIV H(3 2 and the Ha of amino acid i+3 with respect to CysIV (Asp 23 in the present case).…”