Prion diseases are assumed to be caused by the infectious isoform, PrP S c , of a single cellular surface protein, PrP C . PrP S c is an insoluble form of PrP C and is believed to possess a different three-dimensional fold. It may propagate by causing PrP C to adopt its own infectious conformation by an unknown mechanism. Studies on folding and thermodynamic stability of prion proteins are essential for understanding the processes underlying the conversion from PrP C to PrP S c , but have so far been hampered by the low solubility of prion proteins in the absence of detergents. Here, we show that the amino-terminally truncated segment of mouse PrP comprising residues 121 to 231 is an autonomous folding unit. It consists predominantly of α-helical secondary structure and is soluble at high concentrations up to 1 mM in distilled water. PrP(121-231) undergoes a cooperative and completely reversible unfolding/refolding transition in the presence of guanidinium chloride with a free energy of folding of -22 kJ/mol at pH 7. The intrinsic stability of segment 121-231 is not in accordance with present models of the structure of PrP C and PrP S c PrP(121-231) may represent the only part of PrP C with defined three-dimensional structure.