Alkaline Phosphatases : Structure, substrate specificity and functional relatedness to other members of a large superfamily of enzymes

Abstract

Our knowledge of the structure and function of alkaline phosphatases has increased greatly in recent years. The crystal structure of the human placental isozyme has enabled us to probe salient features of the mammalian enzymes that differ from those of the bacterial enzymes. The availability of knockout mice deficient in each of the murine alkaline phosphatase isozymes has also given deep insights into their in vivo role. This has been particularly true for probing the biological role of bone alkaline phosphatase during skeletal mineralization. Due to space constraints this mini-review focuses exclusively on structural and functional features of mammalian alkaline phosphatases as identified by crystallography and probed by site-directed mutagenesis and kinetic analysis. An emphasis is also placed on the substrate specificity of alkaline phosphatases, their catalytic properties as phosphohydrolases as well as phosphodiesterases and their structural and functional relatedness to a large superfamily of enzymes that includes nucleotide pyrophosphatase/phosphodiesterase.

Figure 1

Three-dimensional structure of PLAP. Overview of the structure of human PLAP from the crystallographic coordinates determined by Le Du et al. [4]. Monomer A is shown in ribbon representation and in cyan, while monomer B is shown in surface representation in yellow. Indicated are the active site metals, Zn1, Zn2 and Mg, the novel fourth metal site occupied by Ca, the crown domain and the amino terminal arm. The relative location of the GPI anchor on the processed enzyme is also indicated.

Figure 2

Comparison of the residues coordinating to the active site metals in PLAP and ECAP. The upper panels focus on the environment of the Zn1 and Zn2 metal sites and their ligands while the lower panels display the environment of the Mg metal site and its ligands. Water molecules are shown as red spheres. Green dotted lines denote metal-ligand interactions and hydrogen bonds. The figure is taken from Kozlenkov et al. [6] and is reproduced with permission from the Journal of Biological Chemistry.

Figure 3

Location and environment of the Glu429 and Tyr367 residue. Upper panel ) Top view of the entrance to the active site, showing the position of the Y367 residue from one subunit (wireframe representation) in the immediate vicinity of the Glu429 residue of the other subunit (spacefill representation). Lower panel ) Detail of the active site entrance in spacefill representation showing the location of Glu429 relative to Zn1 and the active site PO₄³⁻ in one subunit and the relative location of Tyr367 of the other subunit. The upper panel was taken from Kozlenkov et al. [6] and is reproduced with permission from the Journal of Biological Chemistry.