X-ray reflectivity from an air-buffer interfacial beta-casein monomolecular film placed on a solution of chymosin (rerun) showed unexpectedly slow proteolytic cleavage. To understand this, the separate structures of beta-casein and chymosin, the presentation of each molecule to the other at the air/liquid interface, and that of their mixtures is reported. At the air/solution interface, the hydrophobicity of the protein molecules causes orientation and some deformation of the conformation. When beta-casein was presented to a chymosin monomolecular interfacial film, the chymosin was largely displaced from the surface, which was accounted for by the different surfactancy of the two molecules at 25 degrees C. There was no observable proteolysis. In the reverse experiment, a significant enzymatic degradation and the signature of hydrophobic fragments was observed but only at and above an enzyme concentration of 0.015 mg/mL in the substrate. For comparison, the air/solution interface of premixed beta-casein with chymosin in phosphate buffer showed that the film was composed of beta-casein proteolytic fragments and chymosin.