Origins of allostery in vertebrate hemoglobin evolution

Abstract

Allostery is an essential and structurally complex form of biochemical regulation, but how allosteric proteins evolved from nonallosteric precursors is unknown. Vertebrate hemoglobin (Hb), a symmetrical tetramer, binds organic phosphates in a central cavity between subunits, which reduces its oxygen affinity. Using ancestral protein reconstruction, we show that Hb's historical precursor, a non-allosteric dimer, was fortuitously on the evolutionary edge of allostery: just three historical substitutions can confer allostery upon it -- one that causes tetramerization, and two others that create an effector-binding site in the cavity. The ancient tetramer could also have acquired inverse allostery -- effector binding that improves oxygen affinity -- via a single substitution that moves the binding site deeper in the cavity. These short evolutionary paths were possible because a key prerequisite for allostery -- propensity to occupy multiple conformations that change upon oxygen binding -- is an intrinsic, ancient property of the globin fold. Evolution of symmetric tetramerization caused this tertiary lability to propagate into oxygen-linked quaternary changes affecting the cavity, so the only remaining requirement for allostery was effector binding. Conformational heterogeneity and multimeric symmetry are widespread, suggesting that many allosteric proteins may have evolved by simple mechanisms from precursors fortuitously poised on the edge of allostery.