Really nice work, revisiting one of biology's paradigms. I have two doubts. I wonder why the experiments shown in Figure 5 compare the t142S single mutant with 4CC. Wouldn´t it be cleaner to compare with Anc1? Second, and it concerns a more general comment: the manuscript focuses on effector allostery, often referred to as heterotrophic allostery. But the paradigm is homotropic allostery, in which an initial O2 binding to a single monomer results in cooperative binding to the three other monomers. This is likely much more complicated to study, but I wonder how heterotrophic and homotrophic allostery relate. Did these evolve separately, or did one follow the other?