The N-terminal 63-residue fragment of the phage 434-repressor, 434(1-63), has a well-defined globular fold in H(2)O solution, and is unfolded in 6 M urea at pH 7.5. In this study, 434(1-63) has been refolded by adding either 1.7 M NaCl or 0.47 M NaTFA to the solution in 6 M urea, and the NMR structures of both refolded forms have been determined. The two refolded forms have similar free energies of unfolding and are approximately 16 kJ/mol less stable than the protein in H(2)O solution. 434(1-63) refolded with NaCl exhibits NMR chemical shifts very similar to and a three-dimensional structure nearly identical to those of 434(1-63) in H(2)O solution. The protein refolded with NaTFA also has a similar global fold, but it shows local differences near Phe44, of which two different orientations of the aromatic ring are compatible with the experimental data. This local conformational polymorphism attracted our interest because hydrophobic contacts between two subdomains of residues 1-36 and 45-63 are mediated by the Phe44 side chain. Anion binding experiments suggest that this polymorphism is caused by binding of TFA(-) anions to a cluster of positively charged Arg and Lys residues located in the loop connecting the two subdomains, with apparent binding constants for TFA(-) (K(app)) on the order of 30 mM(-1).