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Catalytic promiscuity in the alkaline phosphatase superfamily [electronic resource] : identifying specificity determinants in the diesterase family leads to a hypothesis of a "generalist" core bimetallo site found throughout the superfamily

Catalytic promiscuity, the property of enzymes possessing low levels of activity toward non-cognate reactions, can be exploited as a functional tool to investigate conserved and non-conserved mechanism of enzyme specificity and catalysis between members in the same superfamily. Members of the main branch of the Alkaline Phosphatase (AP) superfamily have a structurally conserved core and active site bimetallo site. Other active site features that confer specificity for a given reaction differ between the families. Families within this superfamily catalyze a wide range of reactions, and enzymes within different families show catalytic promiscuity toward reactions catalyzed by other families within this branch. Structural comparisons and phylogenetic analysis suggests that all of the families in the superfamily arose from a common ancestor whose active site consisted of the bimetallo site alone, absent of peripheral, specificity determining features. Experimental data with a member of the nucleotide pyrophosphatase/phosphodiesterase (NPP) family suggests that a "minimal" mutant version of this enzyme that lacks peripheral, specificity determining features, has equal activity toward the two major reactions catalyzed by the AP superfamily, phosphate monoester and phosphate diester hydrolysis reactions. Together, the structural comparisons, phylogenic analysis, and experimental results lead to the hypothesis that the common ancestor of the AP superfamily is a "generalist." By using a variety of techniques, we provide support for a mechanism of evolution in which a "generalist" enzyme may give rise to multiple enzymes specific for, related, but individual reactions. Support for this mechanism, first proposed in 1976, has, until, now been limited. We suggest that this "generalist" mechanism is a valid mechanism for the evolution of ancient enzymes, present in the early evolution of life, into diverse superfamilies in which each family possesses specificity for one given reaction.