The information gain (as measured by the Kullback–Leibler [Kullback and Leibler, 1951] divergence from the SwissProt/TrEMBL [Apweiler et al., 2004] amino acid composition) is then calculated at the corresponding match states within the HMMs extracted above. Next, we interrogate the underlying amino acid probabilities modeled by the most informative HMM and assume that a reduction in the amino acid probabilities (when comparing the wild-type to the mutant residue) indicates a potentially negative impact upon protein function whereas a gain in the amino acid probabilities indicates a more favorable substitution. Furthermore, we assume that larger reductions in amino acid probabilities have more substantial effects than smaller reductions in amino acid probabilities. Here, the predicted magnitude of the effect upon protein function is calculated as follows:
