Allosteric Models for Multimeric Proteins:  Oxygen-Linked Effector Binding in Hemocyanin

Abstract

In many crustaceans, changing concentrations of several low molecular weight compounds modulates hemocyanin oxygen binding, resulting in lower or higher oxygen affinities of the pigment. The nonphysiological effector caffeine and the physiological modulator urate, the latter accumulating in the hemolymph of the lobster Homarus vulgaris during hypoxia, increase hemocyanin oxygen affinity and decrease cooperativity of oxygen binding. To derive a model that describes the mechanism of allosteric interaction between hemocyanin and oxygen in the presence of urate or caffeine, studies of oxygen, urate, and caffeine binding to hemocyanin were performed. Exposure of lobster hemocyanin to various pH values between 7.25 and 8.15 resulted in a decrease of p50. In this pH interval, p50 decreases from 95 to 11 Torr without effectors and from 49 to 6 Torr and from 34 to 5 Torr in the presence of 1 mM urate or caffeine, respectively. Thus, the allosteric effects induced by protons and urate or caffeine are coupled. In contrast, isothermal titration calorimetry did not reveal any differences in binding enthalpy (ΔH°) for urate or caffeine under either normoxic or hypoxic conditions at different pH values. Despite these apparently conflicting results, they can be explained by the nested MWC model if two different types of modulator binding sites are assumed, an allosteric and a nonallosteric type of site. Simulations of in vivo conditions with this model indicate that the naturally occurring modulator urate is physiologically relevant in H. vulgaris only during hypoxic conditions, i.e., either during environmental oxygen limitation or extensive exercise.

Copper Active Sites in Biology

“…The fitting of these models to Hc O 2 affinity for species with a normal Bohr effect indicates that protons stabilize the lower affinity states. 212,214 In the case of organisms with a reverse Bohr effect such as snails, the application of a pH dependent MWC two-state model indicates that protons increase the affinity of either the T-state in H. pomatia 264 or the R-state in L. stagnalis 203 . Data for the horseshoe crab L. polyphemus indicate that in the region of the reverse Bohr effect, protons stabilize the high affinity state decreasing the values of L in a nested model.…”

Lactate accumulation in the intertidal hermit crab, <i>Pagurus samuelis</i>, in response to burial-induced hypoxia

“…If hypoxic conditions persist, concentrations of urate (Dykens, 1991) and the respiratory pigment, hemocyanin (deFur et al, 1990) will increase. Urate allosterically binds to hemocyanin causing a greater affinity for oxygen and increased delivery to the tissues (Menze et al, 2005). In field conditions, if oxygen concentrations are reduced, crustaceans may respond behaviorally by moving in or out of the area (Matabos et al, 2012), engaging in atypical interspecific interactions (Pretterebner et al, 2012), or having reduced ability for handling prey items (Mistri, 2004).…”

The refined structure of functional unit h of keyhole limpet hemocyanin (KLH1‐h) reveals disulfide bridges

“…Between them one molecule oxygen is reversibly bound in side-on (l-g 2 :g 2 ) coordination (4,7,9,10). In most hemocyanins, oxygen binding is highly cooperative and tightly controlled by allosteric effectors (1,2,11,12).…”