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The cellular prion protein of the mouse, mPrPC, consists of 208 amino acids (residues 23–231). It contains a carboxy‐terminal domain, mPrP(121–231), which represents an autonomous folding unit with three α‐helices and a two‐stranded antiparallel β‐sheet. We expressed the complete amino acid sequence of the prion protein, mPrP(23–231), in the cytoplasm of Escherichia coli. mPrP(23–231) was solubilized from inclusion bodies by 8 M urea, oxidatively refolded and purified to homogeneity by conventional chromatographic techniques. Comparison of near‐UV circular dichroism, fluorescence and one‐dimensional 1H‐NMR spectra of mPrP(23–231) and mPrP(121–231) shows that the amino‐terminal segment 23–120, which includes the five characteristic octapeptide repeats, does not contribute measurably to the manifestation of three‐dimensional structure as detected by these techniques, indicating that the residues 121–231 might be the only polypeptide segment of PrPCwith a defined three‐dimensional structure.

The recombinant murine prion protein, mPrP(23–231), was expressed in E. coliwith uniform 15N‐labeling. NMR experiments showed that the previously determined globular three‐dimensional structure of the C‐terminal domain mPrP(121–231) is preserved in the intact protein, and that the N‐terminal polypeptide segment 23–120 is flexibly disordered. This structural information is based on nearly complete sequence‐specific assignments for the backbone amide nitrogens, amide protons and α‐protons of the polypeptide segment of residues 121–231 in mPrP(23–231). Coincidence of corresponding sequential and medium‐range nuclear Overhauser effects (NOE) showed that the helical secondary structures previously identified in mPrP(121–231) are also present in mPrP(23–231), and near‐identity of corresponding amide nitrogen and amide proton chemical shifts indicates that the three‐dimensional fold of mPrP(121–231) is also preserved in the intact protein. The linewidths in heteronuclear 1H–15N correlation spectra and 15N{1H}‐NOEs showed that the well structured residues 126–230 have correlation times of several nanoseconds, as is typical for small globular proteins, whereas correlation times shorter than 1 nanosecond were observed for all residues of mPrP(23–231) outside of this domain.