Metalloproteins and Metallopeptides –Natural Metallofoldamers - Metallofoldamers: Supramolecular Architectures from Helicates to Biomimetics - page 5

Figure 1.3 Stereo view of crystal structure (PDB 1F6S) of bovine holo a-lactalbumin, show-
ing the “Ca 2þ elbow” in lavender. The structure of hen egg white lysozyme (3LZT; yellow) is
superimposed onto the a-lactalbumin structure, showing the significant similarity in folding.
There are four disulfide bonds in a-lactalbumin (Figure 1.3, red), but none in the EF-
hand proteins, which dramatically stabilize the protein conformation [8]. The correct
folding of the protein, which relies on the correct formation of the proper disulfide bonds,
is promoted by the high Ca 2þ -binding affinity to the protein. Ca 2þ binding however does
not change the secondary structure based on circular dichroism (CD) and fluorescence
studies [15,16] which can effectively afford the molten globular state of the protein in the
absence of Ca 2þ possessing a native-like secondary structure but a flexible tertiary struc-
ture [16]. The molten globular state of the protein can be obtained by removal of calcium,
in the presence of denaturants, or in an acid denaturant state [17].
Ca 2þ does not have a significant effect on the metal-binding site, but it does affect the
cleft at the opposite face (underneath the b-sheets shown in Figure 1.3) of the molecule at
the joining of the a-helical (Figure 1.3, light brown) and the b-sheet (Figure 1.3, cyan)
lobes by disturbing the H-bonding pattern in this region, which results in a more open
conformation in apo a-lactalbumin and demonstrates the significance of Ca 2þ in the fold-
ing of a-lactalbumin [18].
Bovine a-lactalbumin (BLA) shares 38% sequence homology with hen egg white lyso-
zyme (HEWL) with most of the differences at charged residues in BLA. However, their
tertiary structures are nearly superimposable with all the four conserved disulfide bonds
(Figure 1.3, yellow ribbon). Electrostatic interactions can stabilize partially unfolded con-
formations,whichcanaidintheformationofmoltenglobulestateinBLA,butnotin
HEWL. The thermodynamic folding barriers in the two proteins are different with a mar-
ginal barrier possible for BLA due to stabilization of partially unfolded conformations in
the presence of Ca 2þ , which stabilizes the fully folded state of the protein [19]. Such elec-
trostatic interactions can be potentially engineered in artificial systems as a means to aid
in structure stabilization. These studies again reveal the significance of metal ions in the
proper folding and stabilization of proteins in order to afford functional natural
metallofoldamers.