Understanding Allosteric Transitions at the Molecular Level

Hemoglobin (Hb) was one of the first protein structures to be solved using X-ray crystallog raphy, and, as it displays positive cooperativity for oxygen binding, has served as the standard for correlating atomic level interactions and allostery (15,16).

Structurally, Hb is a tetrameric protein composed of four subunits—two identical a subunits (referred to as al and a2) and two identical /3 subunits (referred to as /31 and J32). The subunits are arranged such that there are

Figure 9.8. The malate dehydrogenase dimer, indicating the location of the active sites in each protein plus the dimer interface. Malate dehydrogenase demonstrates substrate inhibition that has been attributed to subunit interactions and allosteric regulation by citrate, although the crystal structure of the protein reveals the absence of a separate allosteric site for citrate. See color insert.

Figure 9.8. The malate dehydrogenase dimer, indicating the location of the active sites in each protein plus the dimer interface. Malate dehydrogenase demonstrates substrate inhibition that has been attributed to subunit interactions and allosteric regulation by citrate, although the crystal structure of the protein reveals the absence of a separate allosteric site for citrate. See color insert.

six intersubunit interfaces: ala2, 01/32, alj81, «2/32, al/32, and a2j81. In dilute solutions, the Hb tetramer dissociates into two a/3 dimers: al/31 and ct2/32. Hence this protein is also referred to as a dimer of dimers.

Two structural end states of Hb have been determined through X-ray crystallography—a low affinity deoxy state (tense or T-state) (17, 18) and a high affinity oxy state (relaxed or R-state) (19, 20). Analyses of these two structures indicate that the allosteric transition from the non-liganded deoxy state to the fully liganded oxy state is accompanied by both tertiary and quaternary conformational changes. A15° rotation of the «1/31 dimer with relation to the a2j82 dimer is observed on ligation. This rotation is triggered by structural modification of the heme groups during oxygen binding and is accompanied by numerous changes in noncovalent contacts between residues at the dimer-dimer interface. The T-state dimer-dimer interface possesses more stabilizing noncovalent interactions, and is substantially more stable than the R-state dimer-dimer interface.

Advances in X-ray crystallography and nuclear magnetic resonance spectroscopy have facilitated the solution of a range of other allosteric proteins. These structures have contributed significantly to our understanding of the nature of allostery and the complex atomic level changes that accompany transitions from active/high affinity conformations to in active/low affinity conformations. Examples include: fructose-1,6-bisphosphatase (21), glucos-mine-6-phosphate deaminase (22), chorismate mutase (23), pyruvate kinase (24), hemocyanin (25), d-3-phosphoglycerate dehydrogenase (26), glutarnine-5-phospho-l-pyrophosphatase (27), and lactate dehydrogenase (28).

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