[2] The enzymes from these two sources are 34% homologous, and structural studies have shown that the placement of the catalytic groups in the active sites is virtually identical.

[9] The interface between the two monomers is narrow and well defined, consisting of neutral or apolar amino acids, suggesting the hydrophobic interaction is important for dimerization.

[2] Although the mechanistic steps of the reaction are not disputed, the contributions of various factors to catalysis such as electrostatics, hydrogen bonding of the oxyanion hole, and distal binding effects are discussed below and still debated.

The Warshel group applied statistical mechanical computational methods and empirical valence bond theory to previous experimental data.

[16] The Boxer group used experimental Stark spectroscopy methods to identify the presence of H-bond-mediated electric fields within the KSI active site.

More recently, the Herschlag group utilized unnatural amino acid incorporation to assay the importance of Tyr-14 to KSI catalysis.

There was very little difference in KSI catalytic turnover with decreasing pKa, suggesting, in contrast to the electrostatic studies outlined above, that oxyanion hole stabilization is not primarily important for catalysis.

[23] Sigala et al. found that solvent exclusion and replacement by the remote hydrophobic steroid rings negligibly alter the electrostatic environment within the KSI oxyanion hole.

[24] In addition, ligand binding does not grossly alter the conformations of backbone and side chain groups observed in X-ray structures of PI KSI.

[25][26] Recently, the Herschlag group proposed that remote binding of hydrophobic regions of the substrate to distal portions of the active site contribute to KSI catalysis (>5 kcal/mol).

This activity ratio persists throughout mutagenesis of residues important to oxyanion hole stabilization, implying that distal binding is what accounts for the large aforementioned reactivity difference.

Low-barrier hydrogen bonds and unusual pKa values for the catalytic residues have been proposed as the basis for the fast action of KSI.

[10][15] Gerlt and Gassman proposed the formation of unusually short, strong hydrogen bonds between KSI oxanion hole and the reaction intermediate as a means of catalytic rate enhancement.