Dynamics of allostery in hemoglobin: roles of the penultimate tyrosine H bonds

J Mol Biol. 2006 Feb 17;356(2):335-53. doi: 10.1016/j.jmb.2005.11.006. Epub 2005 Nov 22.


The tyrosine residues adjacent to the C termini of the hemoglobin (Hb) subunits, alphaY140 and betaY145, are expected to play important structural roles, because the C termini are the loci of T-state quaternary salt-bridges, and because the tyrosine side-chains bridge the H and F helices via H bonds to the alphaV93 and betaV98 carbonyl groups. These roles have been investigated via measurements of oxygen binding, (1)H NMR spectra, resonance Raman (RR) spectra, and time-resolved resonance Raman (TR(3)) spectra on site mutants in which the Hcdots, three dots, centeredF H bonds are eliminated by replacing the tyrosine residues with phenylalanine. The TR(3) spectra confirm the hypothesis, based on TR(3) studies of wild-type Hb, that the Hcdots, three dots, centeredF H bonds break and then re-form during the sub-microsecond phase of the R-T quaternary transition. The TR(3) spectra support the inference from other mutational studies that the alphabeta dimers act as single dynamic units in this early phase, motions of the E and F helices being coupled tightly across the dimer interface. Formation of T quaternary contacts occurs at about the same rate in the mutants as in HbA. However, these contacts are weakened substantially by the Y/F substitutions. Equilibrium perturbations are apparent also, especially for the alpha-subunits, in which relaxation of the Fe-His bond, strengthening of the Acdots, three dots, centeredE interhelical H bond, and weakening of the "switch" quaternary contact in deoxyHb are all apparent. Structural effects are less marked for the beta-chain Y/F replacement, but the Bohr effect is reduced by 25%, indicating that the salt-bridge and H bond interactions of the adjacent C terminus are loosened. The alpha-chain replacement reduces the Bohr effect much more, consistent with the global perturbations detected by the structure probes.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Allosteric Regulation
  • Carbon Monoxide / chemistry
  • Hemoglobins / chemistry*
  • Hemoglobins / genetics
  • Hydrogen Bonding*
  • Models, Molecular
  • Nuclear Magnetic Resonance, Biomolecular
  • Oxygen / chemistry
  • Protein Binding
  • Protein Conformation*
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Spectrum Analysis, Raman
  • Tyrosine / chemistry*


  • Hemoglobins
  • Recombinant Proteins
  • Tyrosine
  • Carbon Monoxide
  • Oxygen