Metalloproteins and Metallopeptides –Natural Metallofoldamers - Metallofoldamers: Supramolecular Architectures from Helicates to Biomimetics - page 11

Figure 1.9 Crystal structure of as-isolated dimeric human SOD1 (PDB ID 2C9U). The Cu
ions are shown in blue and Zn ions in red.
6A apart. The coordination site of Cu 2þ is completed by three more His residues and a
water molecule in a square pyramidal geometry, whereas the Zn 2þ tetrahedral site is com-
prised of two more His residues and an Asp residue. The catalytic cycle starts with the
binding of superoxide to Cu 2þ by displacing the coordinated water followed by electron
transfer to the copper and diffusion of oxygen, which results in a trigonal planar Cu þ site.
A second electron transferred by another superoxide results in the regeneration of the
Cu 2þ center and the release of peroxide [57].
SOD1 is properly folded through posttranslational modifications which proceed via two
distinct pathways, depending on whether or not the copper chaperone CCS is required for
the insertion of Cu and the formation of an intramolecular disulfide bond [58]. The forma-
tion of disulfide bridges is crucial in the oligomerization of the protein as the reduced
metal free protein favors the monomeric state. Proper folding of SOD1 is important since
many mutant forms of this protein have been shown to cause amyotrophic lateral sclerosis
(ALS), suggested to be due to destabilized or completely unfolded structures and aggrega-
tion at room temperature [59]. More specifically, the immature reduced forms of the
mutant protein without the formation of disulfide bonds [60] have been implicated in the
aggregation process as they can form incorrect intermolecular disulfide crosslinks. The
spinal cords of ALS transgenic mice have been found to contain significant amounts of
insoluble aggregates composed of such crosslinked multimers, which however are not
observed in other tissues such as the brain cortex and liver [60]. Transgenic mice express-
ing the human mutant G85R SOD1 protein develop paralytic ALS symptoms along with
the appearance of SOD1-enriched inclusions in their neural tissues. The crystal structure
of this mutant supports that metal-deficient and/or disulfide-reduced SOD1 mutants may
contribute to toxicity in SOD1-linked ALS [61].
The unfolding process of SOD1 has been shown to include more than two states,
involving other intermediates in the unfolding process. The irreversible inactivation pro-
cess due to thermal denaturation has distinct features between apo- and holoenzymes,
with the rates of inactivation showing a biphasic response as a function of temperature for
the apoenzyme but a monophasic function in the case of the apoenzyme [62,66]. The role
of the metal ions has been also implicated in the stabilization of the b-barrel structure of
the protein fold [63]. Moreover, the unfolded state of the protein is also stabilized by
metal ions [64]. The derivative Cu, E-SOD1 without a bound Zn 2þ , has a lower thermal
stability, supporting the primary structural role of Zn 2þ in this protein. In addition, Co(II)