Nano gel filtration reveals how fish hemoglobins release oxygen: The Root Effect.

The Root Effect is to many species of fish what the Bohr Effect is to humans regarding the release of O2 from their hemoglobins at low pH. However, Root Effect hemoglobins accomplish this more extensively than human adult hemoglobin in order to satisfy the diverse oxygen requirements in fish. To understand this difference between fish and human hemoglobins, we studied their subunit interface strengths using very low (nanomolar) concentrations, referred to as nano gel filtration. Root Effect hemoglobins in their CO form dissociate in a tetramer-monomer equilibrium. In contrast, tetramers and dimers but no monomers are found for adult human hemoglobin consistent with its well known tetramer-dimer equilibrium. By analogy to the human variant Hb Kansas and a similar recombinant Hb, both of which readily release oxygen due to an unstable oxygenated structure, the mechanism proposed is that oxygenated Root Effect tetramers release their oxygen to form energetically stable deoxygenated tetramers rather than dissociate to energetically unfavorable oxygenated dimers with labile interfaces. In contrast, the strong binding of CO permits observation of dissociation to monomers, thus revealing an intrinsic property of Root Effect fish hemoglobins enabling it to function as an oxygen pump.