||"Before the formation of a complex, the protein and ligand are separately solvated and move freely in the bulk solvent phase. When these two independent 'particles' merge into one, forming the complex, they sacrifice rotational and translational degrees of freedom as two independent particles are reduced to one [ref 28]. This entropic loss is associated with a cost in the Gibbs free energy which has been calculated to be approximately 15–20 kJ per mol [primary source]. Experimentally, this value has been confirmed by Nazare et al. [ref 30] and Borsi et al. [ref 31], who studied the coupling of two non-overlapping fragments binding to factor Xa or matrix metalloproteinase 12 (MMP12)."