Proposed second magnesium binding site. (A) The magnesium
binding
sites in EntC (PDB ID 3HWO). The catalytic magnesium is labeled 1, and the additional
site is labeled 2. Isochorismate is shown in cyan sticks. The general
base (K147) and general acid (E197) are shown in yellow. The residues
of the turn that bind the secondary magnesium are shown in green.
This turn connects an α-helix to the β-strand that is
initiated at the general base. (B) Sequence alignment for the loop
highlighted in (A), comparing PchA, EntC, and Irp9. It should be noted
that the residue side chain (colored) that chelates the second magnesium
in the 3HWO structure
is not conserved among the three proteins. The lysine in bold is the
general base residue. (C) Steady-state magnesium dependence for three
forms of Irp9: WT (circles), V192D (squares), and V192G (triangles).
(D) Steady-state magnesium dependence for three forms of EntC: WT
(circles), D146G (squares), and D146V (triangles). (E) Steady-state
magnesium dependence for three forms of PchA: WT (circles), G220D
(squares), and G220V (triangles).