Reactivity and Dynamics of H₂S, NO, and O₂ Interacting with Hemoglobins from <i>Lucina pectinata</i>

Hemoglobin HbI from the clam <i>Lucina pectinata</i> is involved in H₂S transport, whereas homologous heme protein HbII/III is involved in O₂ metabolism. Despite similar tertiary structures, HbI and HbII/III exhibit very different reactivity toward heme ligands H₂S, O₂, and NO. To investigate this reactivity at the heme level, we measured the dynamics of ligand interaction by time-resolved absorption spectroscopy in the picosecond to nanosecond time range. We demonstrated that H₂S can be photodissociated from both ferric and ferrous HbI. H₂S geminately rebinds to ferric and ferrous out-of-plane iron with time constants (τ_gem) of 12 and 165 ps, respectively, with very different proportions of photodissociated H₂S exiting the protein (24% in ferric and 80% in ferrous HbI). The Gln­(E7)­His mutation considerably changes H₂S dynamics in ferric HbI, indicating the role of Gln­(E7) in controling H₂S reactivity. In ferric HbI, the rate of diffusion of H₂S from the solvent into the heme pocket (<i>k</i>_entry) is 0.30 μM^–1 s^–1. For the HbII/III–O₂ complex, we observed mainly a six-coordinate vibrationally excited heme–O₂ complex with O₂ still bound to the iron. This explains the low yield of O₂ photodissociation and low <i>k</i>_off from HbII/III, compared with those of HbI and Mb. Both isoforms behave very differently with regard to NO and O₂ dynamics. Whereas the amplitude of geminate rebinding of O₂ to HbI (38.5%) is similar to that of myoglobin (34.5%) in spite of different distal heme sites, it appears to be much larger for HbII/III (77%). The distal Tyr­(B10) side chain present in HbII/III increases the energy barrier for ligand escape and participates in the stabilization of bound O₂ and NO