306 GROUP I AND II METALS IN BIOLOGICAL SYSTEMS: GROUP II
The X - ray crystallographic structure of human calmodulin, refi ning to 1.7 - Å
resolution, was obtained in 1992 (PDB: 1CLL).^72 The interhelical angle
data are included in Table 6.7. The X - ray crystallographic structure of apo -
calmodulin — one in which calcium ions are not present — was carried out by
Schumacher et al. (PDB: 1QX5).^73 As can be seen from Figure 6.22B , the apo
structure consists of more compact coiled globular domains, whereas the
calcium - saturated structure (PDB: 1EXR) shows the characteristic calmodulin
dumbbell shape. The coloring in Figure 6.22B is the same as that noted above
for Figure 6.22A. In the apo - calmodulin structure (PDB: 1QX5), the central
E and F helices are not contiguous, helices are shorter, and the loop regions
between helices are longer. Note that for the holo - calmodulin structure (PDB:
1EXR), helices A and B (and helices C and D) have opened up relative to
each other, forming the hydrophobic cleft where drug molecules or incoming
target enzymes can bind.
The solution structure of Ca 2+ - free calmodulin has been determined
for several species. In 1995, Zhang, Tanaka, and Ikura published the apo -
calmodulin NMR structure for Xenopus leavis (PDB: 1DMO).^74 Another apo -
calmodulin NMR structure for Xenopus leavis was published in the same year
by the Ad Bax group (PDB: 1CFC, 25 structures, and 1CFD, averaged struc-
ture).^75 These researchers used three - and four - dimensional heteronuclear
NMR experiments, including ROE, isotope - fi ltering combined with reverse
labeling, and measurement of more than 700 three - bond J - couplings. (See
Sections 3.4.3 , 3.4.10 , and 3.4.11 for descriptions of these NMR techniques.)
As is true for the PDB: 1DMO structure, the PDB: 1CFD protein consists of
two globular domains separated by a fl exible linker, with no stable, direct
contacts between the two domains. In the absence of Ca 2+ , the four helices in
each of the two globular domains form a highly twisted bundle, each capped
by a short antiparallelβ - sheet. (This arrangement is qualitatively similar to
that observed in the crystal structure of the Ca 2+ - free N - terminal domain of
troponin C.) Other research groups have determined solution structures of
apo - calmodulin.^76 In all cases, removal of Ca 2+ ions from the structure changes
the conformation of calmodulin to close the hydrophobic pockets that would
bind to target enzymes.
More recently, workers in Japan published the solution structure of yeast
(Saccharomyces cerevisiae ) apo - calmodulin (PDB: 1LKJ).^77 Yeast calmodulin
is 60% identical in its amino acid sequence with vertebrate CaMs. The 1LKJ
N - terminal domain with its two helix – loop – helix calcium - binding domains
looks quite similar to those of 1DMO and 1CFD (see Figure 6.23 ).
The central linker region is more or less unwound and different for each of
the three apo - calmodulins discussed here and shown in Figure 6.23. In addi-
tion, the C - terminal region takes different conformations in each structure.
While the three illustrated C - terminal sections still each consist of two basic
helix – loop – helix calcium - binding domains, in (yeast) PDB: 1LKJ the segment
corresponding to the fourth Ca 2+ - binding site deviates in its primary structure
from a typical EF - hand motif and loses the ability to bind calcium ion. Note