Probing protein interaction chemistry through crystal growth: structure, mutation, and mechanism in subtilisin s88

Observations of systematic variation in the shapes of protein crystals have unique potential to report chemical effects on protein-protein interaction s, because diffraction can be used to image the detailed structure that lin ks an easily controlled "cause" (change in chemical conditions) with an eas ily observed "effect" (change in crystal shape). Crystal shape correlates d irectly with the relative growth rates of the various faces. By studying ca ses of known structures in which crystal shape varies systematically with s ome solution parameter (pH, ionic strength, concentration of a specific sol ute, etc.), it is sometimes possible to correlate that parameter with its e ffect on a particular crystal contact. Such correlations provide the basis for models of how solution parameters affect protein-protein interactions. We have applied this technique in a model system involving a subtilisin BPN ' variant. Crystal shape varies systematically with pH and ionic strength, an effect that has been traced to a specific interaction in one crystal con tact. Mutation of an Asp sidechain in this contact (to Asn) eliminates the morphology variation effect, confirming the key role of this residue in the observed shape effect, and enabling the mechanism to be placed in a mathem atical framework. (C) 2000 Elsevier Science B.V. All rights reserved.