Mp. Golinelli et al., EXTENSIVE LIGAND REARRANGEMENTS AROUND THE [2FE-2S] CLUSTER OF CLOSTRIDIUM-PASTEURIANUM FERREDOXIN, Biochemistry, 37(29), 1998, pp. 10429-10437
The [2Fe-2S] cluster of the ferredoxin from Clostridium pasteurianum i
s coordinated by cysteines 11, 56, and 60 and by a fourth cysteine, re
sidue 24 in the wild-type protein, located on a flexible and deletable
loop around residues 14-30. New mutated forms of this ferredoxin show
that the fourth cysteine ligand can be located in any one of position
s 14, 16, 21, 24, or 26. Another set of molecular variants has unveile
d a new case of ligand swapping on the cysteine 60 ligand site. Replac
ement of cysteine 60 by alanine and introduction of a cysteine in posi
tion 21 yielded a ferredoxin that assembles a [2Fe-2S] cluster of whic
h the ligands are cysteines 11, 21, 24, and 56. This cysteine ligand p
attern is similar to that occurring in plant-type or mammalian-type fe
rredoxins, although the overall sequence similarities are below detect
ion. Moreover, the vibrational and electronic properties of the result
ing [2Fe-2S](2+/+) center, as revealed by resonance Raman and EPR stud
ies, are strikingly similar to those of mammalian-type ferredoxins. Th
e extensive set of mutated forms of the C. pasteurianum ferredoxin now
available indicates that cysteine ligand exchange may occur on residu
es 24 and 60, but not on residues 11 and 56. It is thus suggested that
cysteines 24 and 60 are part of a solvent accessible aspect of the Fe
-S cluster, whereas cysteines 11 and 56 are buried and form the more r
igid part of the polypeptide ligand framework. Tn view of the unpreced
ented versatility of this [2Fe-2S] cluster and of its polypeptidic env
ironment, the introduction of ligands other than cysteine in various p
ositions has been attempted. These experiments have remained unsuccess
ful, and even including previous studies, noncysteinyl ligation has be
en obtained with this protein in only very few cases. The data provide
an extensive confirmation that Fe-S clusters have a strong preference
for thiolate ligation and rationalize the relatively rare occurrence
of noncysteinyl ligation in native Fe-S proteins.