The results of a study to measure the beta-sheet forming propensities of the 20 naturally occurring amino acids are presented. The protein host for these studies is the 56 amino acid B1 domain of staphylococcal IgG binding protein G [Fahnestock, S.R., Alexander, P., Nagle, J., & Filpula, D. (1986) J. Bacteriol. 167, 870-880]. This protein was selected because it exhibits a reversible two-state thermal denaturation transition and its structure is known at high resolution. A suitable guest position in the protein was identified, and its neighboring environment was modified to minimize the potential for artifactual interactions. All 20 amino acids were individually substituted at the guest site, and their effect on the protein's thermal stability was determined. NMR was used to verify the structural integrity of several of the proteins with different amino acid substitutions at the guest site. The results of these studies provide a thermodynamic scale for the relative beta-sheet forming propensities of the amino acids that shows a clear correlation with the beta-sheet preferences derived from statistical surveys of proteins of known structure.