Nucleoside diphosphate kinase reversibly transfers the gamma-phosphate of ATP
onto its active site histidine. We have investigated the transition state of
histidine phosphorylation with the high-resolution crystal structures of the
enzyme from Dictyostelium discoideum with MgADP and either aluminium or
beryllium fluoride. The bound aluminium fluoride species is the neutral species
AlF3 and not the more common AlF4-. AlF3 forms a trigonal bipyramid that makes
it an accurate analog of the transition state of the gamma-phosphate of ATP
undergoing transfer to the catalytic histidine. Its axial ligands are a
histidine nitrogen and a beta-phosphate oxygen. Beryllium fluoride also binds at
the same position and with the same ligands but in a tetrahedral geometry
resembling the Michaelis complex rather than the transition state. The two x-ray
structures show explicit enzyme-substrate interactions that discriminate between
the ground and the transition states of the reaction. They also illustrate the
partially dissociative geometry of the transition state of phosphoryl transfer
and demonstrate the potential applications of metallofluorides for the study of
kinase mechanisms.
Figure 1.
Fig. 1. Simulated annealing F[o]-F[c] omit maps contoured at
3.5 showing
ADP (to the left) and His-122 (to the right). (A) AlF[3] omit
map at 2.0-Å resolution. Al3+ (green) in trigonal
bipyramidal configuration is shown coordinating to three
equatorial fluorines (orange) and with partial bonds to ADP O[7]
and to His-122 N . Mg2+
(white) has an octahedral geometry, with a tridendate
coordination to the - and -phosphates
and a fluorine atom and three water molecules (red) as
additional ligands. (B) BeF[3]^ omit
map at 2.3-Å resolution. Be^2+ (green) is modeled in a
tetrahedral geometry, with three fluorines and O[7] as the
fourth ligand. An alternative configuration for Be^2+ has
His-122 N as the
fourth ligand.
Figure 2.
Fig. 2. Schematic representation of NDPK-nucleotide
interactions. (A) ADP-AlF[3] from the x-ray structure. (B)
ADP-BeF[3]^ with
Be^2+ modeled at the -phosphate
O[7] as in Fig. 1B. In all cases, there^ is a short hydrogen
bond between the 3 hydroxyl of
the ribose^ and the -phosphate
O[7].
showing
ADP (to the left) and His-122 (to the right). (A) AlF[3] omit
map at 2.0-Å resolution. Al3+ (green) in trigonal
bipyramidal configuration is shown coordinating to three
equatorial fluorines (orange) and with partial bonds to ADP O[7]
and to His-122 N 
. Mg2+
(white) has an octahedral geometry, with a tridendate
coordination to the 
- and 
-phosphates
and a fluorine atom and three water molecules (red) as
additional ligands. (B) BeF[3]^ 
omit
map at 2.3-Å resolution. Be^2+ (green) is modeled in a
tetrahedral geometry, with three fluorines and O[7] as the
fourth ligand. An alternative configuration for Be^2+ has
His-122 N 
hydroxyl of
the ribose^ and the 
