Abstract
Biological signal transduction commonly involves cooperative interactions in the binding of ligands to their receptors. In many cases, ligand concentrations in vivo are close to the value of the dissociation constant of their receptors, resulting in the phenomenon of ligand depletion. Using examples based on rotational bias of bacterial flagellar motors and calcium binding to mammalian calmodulin, we show that ligand depletion diminishes cooperativity and broadens the dynamic range of sensitivity to the signaling ligand. As a result, the same signal transducer responds to different ranges of signal with various degrees of cooperativity according to its effective cellular concentration. Hence, results from in vitro dose-response analyses cannot be applied directly to understand signaling in vivo. Moreover, the receptor concentration is revealed to be a key element in controlling signal transduction and we propose that its modulation constitutes a new way of controlling sensitivity to signals. In addition, through an analysis of the allosteric enzyme aspartate transcarbamylase, we demonstrate that the classical Hill coefficient is not appropriate for characterizing the change in conformational state upon ligand binding to an oligomeric protein (equivalent to a dose-response curve), because it ignores the cooperativity of the conformational change for the corresponding equivalent monomers, which are generally characterized by a Hill coefficient . Therefore, we propose a new index of cooperativity based on the comparison of the properties of oligomers and their equivalent monomers.
Highlights
Dose-response is one of the most common experimental approaches used by biologists to monitor the properties of signaling molecules
We illustrate the importance of considering ligand depletion with the highly cooperative E. coli flagellar motor system [30], which controls the direction of flagellar rotation in response to the concentration of phosphorylated CheY [31]
The motor bias reflects a change of rotation from counter-clockwise to clockwise and a change of fractional activation, which is influenced by the interaction of CheY-P with the 34 units of FliM comprising the motor ring [32]
Summary
Dose-response is one of the most common experimental approaches used by biologists to monitor the properties of signaling molecules The power of this approach arises from the fact that the change in any quantifiable physiological response can be measured as a function of the chemical stimulus responsible. Because the R conformation has a higher affinity than the T state for a ligand specific for the protein under consideration, the presence of ligand pulls the T{R equilibrium towards the R state Under these conditions, a clear distinction can be made between two mathematical functions that describe the behavior of proteinligand interactions as a function of ligand concentration: 1) the binding function, Y , defined as the fractional occupancy of the ligand binding sites of the protein, taking into account both the T and R states; and 2) the state function, R , defined as the fraction of molecules in the R state.
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