310 GROUP I AND II METALS IN BIOLOGICAL SYSTEMS: GROUP II
binding sites for TFP.^81 The question arises then as to which calmodulin hydro-
phobic pockets bind the drug molecules. Possibilities include: (1) both in the
N - terminal domain; (2) both in the C - terminal domain; (3) one each in the N -
and C - terminal domain; or (4) one in the N - or C - terminal domain and one
in an interdomain region. A series of crystal and solution structural studies
have answered this question. In a 1994 study by Cook and co - workers the
crystal structure of the 1 : 1 CaM - TFP complex (CaM - 1TFP) shows that one
TFP molecule binds in the hydrophobic pocket of the C - domain (PDB:
1CTR).^82 In this crystallographic structure, refi ned to 2.45 - Å resolution, resi-
dues 75 – 80 (midpoint of the central helix) and the terminal residue lys148
show almost no electron density and do not appear in the fi nal map. As with
many other reported calmodulin structures, the central area of the central
helix appears severely bent, disordered, and/or fl exible. In spite of the disor-
dered residues, it can be determined that the compact globular structure of
the PDB: 1CTR, 1 : 1 CaM:TFP complex contrasts markedly with the extended
dumbbell shape exhibited by the X - ray structures of Ca 2+ - saturated calmodu-
lin such as PDB: 3CLN or PDB: 1EXR (see Figures 6.21 and 6.22A ). Thirteen
calmodulin amino acid residues form 191 contacts at 5 Å or less between CaM
and TFP. The TFP tricyclic phenothiazine ring system lies in the same hydro-
phobic binding pocket utilized in binding peptides (peptide binding to be dis-
cussed below) and makes contact with phe92, ile100, leu105, met109, met124
(25 contacts), ile125, glu127, ala128, val136, phe141, and met144 (20 contacts).
The trifl uoromethyl group ( — CF 3 ) interacts with glu11 (the only residue from
the N - terminal region involved in binding TFP), glu123, met124, and glu127.
TFP ’ s aliphatic chain — more interactions are possible if this chain is at least
three carbons long — interacts with met124, glu127, and met144 while the
piperazine ring, which has a net positive charge, interacts with glu127, ala128,
and met144.
In 1998, Ov á di and co - workers studied the structure of calmodulin in
complexes with two TFP molecules as well as complexes with TFP and two
Figure 6.24 The calmodulin antagonist trifl uoperazine, TFP.SN(CH 2 ) 3NNCH 3C
FF
FTrifluoperazine, TFPphenothiazine
ring system