Crystals of haemoglobin with the T quaternary structure bind oxygen noncooperatively with no Bohr effect

Open Access
Publisher Website
Google Scholar
Abstract
THE relationship between the structure and function of haemoglobin has mainly been studied by comparing its X-ray crystal structures with its function in solutions1–11. To make a direct comparison we have studied the functional properties of haemoglobin in single crystals, an approach that has been an important part of the investigation of several enzyme mechanisms12,13. Here we report on the oxygen binding by single crystals of human haemoglobin grown in solutions of polyethylene glycol. Unlike haemoglobin crystals formed in concentrated salt solution, which crack and become disordered on oxygenation14–16, crystals grown in polyethylene glycol remain intact. X-ray studies have shown that the T (deoxy) quaternary structure of haemoglobin in this crystal at pH 7.0 is maintained at atmospheric oxygen pressure, and that the salt-bridges are not broken117–19. We find striking differences between oxygen binding by haemoglobin in this crystal and by haemoglobin in solution. Not only is oxygenation of the crystal noncooperative, but the oxygen affinity is independent of pH in the range 6.0–8.5, and is much lower than that of the T state in solution. The lack of cooperativity without a change in quaternary structure is predicted by the two-state allosteric model of Monod, Wyman and Changeux1,5. The absence of a Bohr effect without breakage of salt-bridges is predicted by Perutz's stereochemical mechanism2,4,9–11. In contrast to the X-ray result that oxygen binds only to the α haems, our measurements show that the α haems have only a slightly higher affinity than the β haems.