Two distinct heme distal site states define Cerebratulus lacteus mini-hemoglobin oxygen affinity

The nerve tissue hemoglobin of Cerebratulus lacteus (CerHb) is the smallest naturally occurring known hemoglobin. Stabilization of the diatomic bound species (e.g., O-2) is achieved through a network of hydrogen bonds based on three key residues TyrB10, GlnE7, and ThrE11. The first two residues are typically associated in hemoglobins with enhanced O-2 affinity, related to hydrogen bond stabilization of the heme-bound O-2 resulting in a decrease of the ligand dissociation rates. In contrast to the above observations, the affinity of CerHb for O-2 is only moderate, and the rate of O-2 dissociation is unexpectedly high. To gain insight on the diverse molecular mechanisms controlling ligand affinities, we have analyzed w.t. CerHb and its ThrE11 -> Val mutant by means of joint molecular dynamics and quantum mechanics simulation techniques, complementing recent site-directed mutagenesis experiments. Our results suggest that the observed O-2 dissociation rates can only be explained through a dynamic equilibrium between high and low affinity states of the w.t. CerHb heme distal site.