The successful incorporation of difluoromethionine (DFM), a novel F-19 NMR probe of internal amino acid packing, into the three methionine positions (1, 14, and 107) of a recombinant protein, the lysozyme from bacteriophage lambda (LaL), is reported. The anisochronous F-19 NMR signals of the diastereotopic fluorines showed a variation in the degree of chemical shift difference when present at relatively free surface positions (Met1 and Met107) versus the tightly packed protein core (Met14), with the anisochronicity greatly enhanced for DFM incorporated at this latter position. The increased magnetic nonequivalence of the two fluorines at position 14 is thought to be a consequence of the restricted environment of DFM at this position. The anisochronicity of these two fluorines is further manifested in a differential chemical shift change for these two fluorines upon binding of an oligosaccharide inhibitor to LaL, with one of the two fluorines experiencing a significant upfield shift compared to the other. This differential variation is thought to be associated with a very subtle change in the protein conformation surrounding one fluorine at position 14, which is not significantly translated to the environment of the other fluorine.