The two-enzyme test model. The first enzyme, E1, binds the initial substrate, S1, to form the enzyme-substrate complex, E1S1. Within this complex, E1 transforms S1 into its product, S2, resulting in the transformation of E1S1 into E1S2, and then E1S2 liberates S2, thus regenerating E1. In a similar manner, the second enzyme E2 binds S2; transforms S2 into S3 and finally releases the final product, S3. The k'if and k'ir are the forward and reverse rate constants; their numbering, i, has been chosen in such a way as to be consistent with that of previous publications.