HETERONUCLEAR RELAXATION STUDY OF THE PH DOMAIN OF BETA-SPECTRIN - RESTRICTION OF LOOP MOTIONS UPON BINDING INOSITOL TRISPHOSPHATE

The structural dynamics of protein ligand binding sites is one factor determining the specificity towards related ligands. In this context, the spectrin PH domain, which binds to a number of phosphatidylinosito l lipid head groups, was investigated with respect to the dynamics of the binding loops. The latter were found to be of intermediate flexibi lity on a picosecond to nanosecond time-scale in the free protein and become more rigid upon ligand binding. Significant N-15 and proton che mical shift changes occur in the binding loops. The internal correlati on time, determined from 15N heteronuclear relaxation data using the s tandard model-free approach, decreases upon ligand binding. For severa l residues a concomitant rise in the generalized order parameter is ob served. This is interpreted as a dampening effect of the ligand on a s low loop motion, while a fast component is not affected. Molecular dyn amics simulations were performed to further investigate this situation . In fact, two timescales of loop motions in the free state are observ ed in a 9 ns molecular dynamics trajectory. Agreement with generalized order parameters obtained from the experiment improves when a subtraj ectory is analyzed that excludes rare dihedral transitions. (C) 1998 A cademic Press.