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M A Daugherty

Publications and source records attributed to M A Daugherty.

4 recordsLinked to original sources
PubMed1994

Bohr effects of the partially-ligated (CN-met) intermediates of hemoglobin as probed by quaternary assembly.

Free energies of quaternary assembly (dimers to tetramers) were determined for the 10 ligation species of CN-methemoglobin in the region of the alkaline Bohr effect (pH 7.0-9.5). Analysis of this database yielded the following principal findings: (1) At each pH, the nine CN-met species exhibit two distinct values of Bohr proton release and Bohr free energy. The two Bohr effects are found to distribute in a fashion that coincides with predictions of a symmetry rule (Ackers et al., 1992), i.e., the first value reflects a "tertiary Bohr effect" arising from ligation within the quaternary T tetramer and a second Bohr effect arises from the quaternary transition (T-->R) which occurs when both dimeric half-molecules acquire at least one ligated subunit. (2) The Bohr effects for CN-met ligation are in good agreement with previously-established Bohr effects for stepwise O2 binding under identical conditions (Chu et al., 1984). (3) In combination with recent studies which show that CN-met species [21] has a quaternary T structure (Daugherty et al., 1991; Doyle & Ackers, 1992; LiCata et al., 1993), the present results show that the "tertiary Bohr effect" within quaternary T exceeds the Bohr effect of dissociated dimers, as suggested by Lee and Karplus (1983). (4) The tertiary Bohr effect is found to account for the pH dependence of tertiary constraint energy, delta Gtc, which "pays" for ligand-binding cooperativity prior to the quaternary (T-->R) switchover. Possible origins of the tertiary Bohr effect and its relationship to the quaternary Bohr effect are considered.

Allosteric Regulation↗
PubMed1992

Molecular code for cooperativity in hemoglobin.

Although tetrameric hemoglobin has been studied extensively as a prototype for understanding mechanisms of allosteric regulation, the functional and structural properties of its eight intermediate ligation forms have remained elusive. Recent experiments on the energetics of cooperativity of these intermediates, along with assignments of their quaternary structures, have revealed that the allosteric mechanism is controlled by a previously unrecognized symmetry feature: quaternary switching from form T to form R occurs whenever heme-site binding creates a tetramer with at least one ligated subunit on each dimeric half-molecule. This "symmetry rule" translates the configurational isomers of heme-site ligation into six observed switchpoints of quaternary transition. Cooperativity arises from both "concerted" quaternary switching and "sequential" modulation of binding within each quaternary form, T and R. Binding affinity is regulated through a hierarchical code of tertiary-quaternary coupling that includes the classical allosteric models as limiting cases.

Allosteric Regulation↗
PubMed1991

Identification of the intermediate allosteric species in human hemoglobin reveals a molecular code for cooperative switching.

The 10 ligation species of human cyanomethemoglobin were previously found to distribute into three discrete cooperative free energy levels according to a combinatorial code (i.e., dependent on both the number and configuration of ligated subunits). Analysis of this distribution showed that the hemoglobin tetramer occupies a third allosteric state in addition to those of the unligated (T) and fully ligated (R) species. To determine the nature of the intermediate allosteric state, we have studied the effects of pH, temperature, and single-site mutations on its free energy of quaternary assembly, in parallel with corresponding data on the deoxy (T) and fully ligated (R) species. Results indicate that the intermediate allosteric tetramer has the deoxy (T) quaternary structure. This finding, together with the resolved energetic distribution of the 10 microstates reveals a symmetry rule for quaternary switching--i.e., switching from T to R occurs whenever a binding step creates a tetramer with one or more ligated subunits on each side of the alpha 1 beta 2 intersubunit contact. These studies also reveal significant cooperativity within each alpha 1 beta 1 dimer of the T-state tetramer. The ligand-induced tertiary free energy alters binding affinity within the T structure by 170-fold prior to quaternary switching.

Allosteric Site↗