The deletion of amino acids 114-121 in the TM1 domain of mouse prion protein stabilizes its conformation but does not affect the overall structure

Abstract

A mutant of mouse prion protein (PrPC) carrying a deletion of residues 114 121 (PrPΔ 114 121) has previously been described to lack convertibility into the scrapie-associated isoform of PrP (PrPSc) and to exhibit a dominant-negative effect on the conversion of wild-type PrPC into PrPSc in living cells. Here we report the characterization of recombinantly expressed PrPΔ 114 121 by Fourier-transformation infrared spectroscopy (FTIR) and circular dichroism (CD) spectroscopy. The analysis of spectra revealed an increased antiparallel β-sheet content in the deletion mutant compared to wild-type PrPC. This additional short β-sheet stabilized the fold of the mutant protein by ΔΔG0=3.4±0.3 kJ mol -1 as shown by chemical unfolding experiments using guanidine hydrochloride. Secondary structure predictions suggest that the additional β-sheet in PrPΔ 114 121 is close to the antiparallel β-sheet in PrPC. The high-affinity Cu2+-binding site outside the octarepeats, which is located close to the deletion and involves His110 as a ligand, was not affected, as detected by electron paramagnetic resonance (EPR) spectroscopy, suggesting that Cu2+ binding does not contribute to the protection of PrPΔ 114 121 from conversion into PrPSc. We propose that the deletion of residues 114 121 stabilizes the mutant protein. This stabilization most likely does not obstruct the interaction of PrPΔ 114 121 with PrPSc but represents an energy barrier that blocks the conversion of PrPΔ 114 121 into PrPSc.