Structural and dynamic characterization of neurogranin and other proteins

Abstract

Full-length rat neurogranin has been investigated by NMR. Though neurogranin is unfolded as evidenced by high backbone mobility and absence of long-range NOE, residual secondary structure in a region of G25-A42 is identified by using various NMR parameters. 15N relaxation data indicates motional restrictions of neurogranin in a region of D15-K48 on nanosecond time scale. Spectral densities and order parameter data further confirm that the unfolded neurogranin exists in a conformation with residual secondary structure. Nascent helical structure of nerogranin may facilitate its binding to target protein like calmodulin. However, the complex between neurogranin and calmodulin is not stable enough for structural determination by NMR. Calmodulin titration of neurogranin indicates that neurogranin residues D15-G52 undergo significant structural changes upon binding to calmodulin. Besides neurogranin characterization, HSQC-TOCSY and HSQC-NOESY spectrum analysis of calmodulin is summarized. Expression and purification of recombinant calmodulin and mPRL proteins are also discussed.