Accurate quantitative measurement of structural dispersion in proteins remains a prime challenge to both X-ray crystallography and NMR spectroscopy. Here we use a model-free approach based on measurement of many residual dipolar couplings (RDCs) in differentially orienting aqueous liquid crystalline solutions to obtain the side chain chi 1 distribution sampled by each residue in solution. Applied to the small well-ordered model protein GB3, our approach reveals that the RDC data are compatible with a single narrow distribution of side chain chi 1 angles for only about 40% of the residues. For more than half of the residues, populations greater than 10% for a second rotamer are observed, and four residues require sampling of three rotameric states to fit the RDC data. In virtually all cases, sampled chi 1 values are found to center closely around ideal g , g(+) and t rotameric angles, even though no rotamer restraint is used when deriving the sampled angles. The root-mean-square difference between experimental (3)J(H alpha H beta) couplings and those predicted by the Haasnoot-parametrized, motion-adjusted Karplus equation reduces from 2.05 to 0.75 Hz when using the new rotamer analysis instead of the 1.1-a X-ray structure as input for the dihedral angles. A comparison between observed and predicted (3)J(H alpha H beta) values suggests that the root-mean-square amplitude of chi 1 angle fluctuations within a given rotamer well is ca. 20. The quantitatively defined side chain rotamer equilibria obtained from our study set new benchmarks for evaluating improved molecular dynamics force fields, and also will enable further development of quantitative relations between side chain chemical shift and structure.