We tested the fractions for their ability to interact with PDZ2 of PSD-95 using the same chemical shift perturbation approach.
Following isolation and chemical characterization, we tested each compound for its ability to bind to PSD-95 PDZ2.
We quantified the binding affinity of the compounds to PDZ2 of PSD-95 by normalizing the chemical shift of the amide groups of S19 and A77 in relation to the maximal shift of these groups upon incubation with the crude extract (100%).
Aqueous extracts of Chinese herbs commonly used in stroke therapy were screened by NMR spectroscopy based on the hypothesis that they might contain small compounds that bind to PDZ2 of PSD-95.
PDZ2 of PSD-95 is believed to be, overall, a quite rigid molecule with a mostly rigid binding groove, with exception of the GLGF-motif-containing loop and the [alpha]B/[beta]F loop, which display limited flexibility in the absence of its targets (Tochio et al.
This perturbation profile suggests that the interaction between the compounds and PDZ2 of PSD-95 largely occurred in the center of the binding groove, and the carboxyl-binding loop and the hydrophobic pocket accommodating the side chain of the extreme C-terminal residue of peptide ligands are unoccupied.
Brenman JE, Chao DS, Gee SH, McGee AW, Craven SE, Santillano DR, Wu Z, Huang F, Xia H, Peters MF, Froehner SC, Bredt DS (1996) Interaction of nitric oxide synthase with the postsynaptic density protein PSD-95 and alphalsyntrophin mediated by PDZ domains.
Christopherson KS, Hillier BJ, Lim WA, Bredt DS (1999) PSD-95 assembles a ternary complex with the N-methyl-D-aspartic acid receptor and a bivalent neuronal NO synthase PDZ domain.