The concepts of stoichiometric and site binding constants

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The interaction between a monovalent ligand L and a multivalent receptor R involves the presence of various species, including the complex of R fully saturated with a number of ligands, and intermediate complexes of R partially saturated. This scenario can be described in terms of reaction schemes following two approaches:


  1. Based on equilibria between existing stoichiometric species (Stoichiometric approach).
  2. Based on equilibria between L and specific interaction sites of R (independent sites approach).
For a better understanding, let´s consider a particular case where L binds to a bivalent receptor:

1. Stoichiometric approach

This approach uses reaction schemes based on equilibria between stoichiometric species and yields stoichiometric binding constants. A model based on stoichiometric equilibria is valid to fit data of both independent and non-independent events and therefore, it is of wider applicability.

Here, the reaction scheme includes a first equilibrium between the free species and the intermediate RL and the second equilibrium between RL + L and RL2 (Fig. 1). The corresponding binding constants, K1 and K2, are denominated stoichiometric binding constants since they refer to equilibria between stoichiometric species.

2. Independent site approach

This approach uses a reaction scheme based on the binding of the ligand to individual sites present in the receptor and considering that all the sites are independent; thus, it supplies site binding constants.

In this case, the reaction scheme considers the presence of two sites in the bivalent receptor and two intermediate complexes (R, L and RL) formed when the ligand binds to s1 or s2 and consequently, the existence of a total of 4 equilibria (Fig. 2). The corresponding binding constants, Ks1, Ks2, Ks1s2 and Ks2s1, are denominated site binding constants since they refer to equilibria between L each specific site of R.

If you want to know more about how to get the stoichiometry (number of sites) and site binding constants with the independent sites approach you can click on the following button:


Eva Muñoz

Dr Muñoz has worked as a researcher for over 15 years in different international centers in Spain, Germany and the USA, focusing on the characterization of complex biomolecular interactions by different experimental techniques such as NMR, FT-IR, fluorescence and Surface Plasmon Resonance (SPR). She has a high publication record in this area as well as two patents on biotechnological developments. Eva is working with us in the validation and design of AFFINImeter as well as in the implementation of functionalities for the analysis of SPR measurements.