Purification and spectroscopic characterization of the recombinant BG21 isoform of murine Golli myelin basic protein

A recombinant form of the murine Golli-myelin basic protein (MBIP) isoform BG21 (rmBG21) has been expressed in E coli, and isolated to 96% purity via metal chelation chromatography. Characteristic yields were 68 mg protein per liter of culture in either minimal M9 or standard Luria-Bertani media. Circular dichroism spectroscopy showed that rmBG21 had a large proportion of random coil in aqueous solution, but gained alpha-helix in the presence of monosialoganglioside Gm, and PI(4)P, as well as in the membrane-mimetic solvent trifluoroethanol. Bioinformatics analyses of the amino acid sequence of rmBG21 predicted an N-terminal calmodulin (CaM)-binding site. It was determined by fluorescence spectroscopy and dynamic light scattering that rmBG21 and CaM interacted weakly in a 1:1 ratio in a Ca2+-dependent manner. Solution NMR spectra of uniformly [(CN)-C-13-N-15]-labeled protein in aqueous buffer were consistent with it being an extended protein; spectral quality was independent of temperature. Thus, like "classic" MBP and the Golli-MBP isoform J37, rmBG21 is intrinsically disordered, implying multi functionality, and that its conformation depends on its environment and bound ligands. (c) 2006Wiley-Liss, Inc.