Identification of Mono-iodotyros1ne from Iodinated Pepsin By

In an earlier paper (1) the writer reported the isolation of/-mono-iodotyrosine from an alkaline hydrolysate of very slightly iodinated pepsin. Since then Harington and Pitt Rivers (2) have synthesized/-mono-iodotyrosine and dlmono-iodotyrosine and a comparison of the properties of their compounds with those reported by us led them to conclude that for the compound from pepsin "the nature remains in doubt." Differences were noted by them in the following properties: 1. Specific Optical Rotation.--The [a]tp~ cent of synthetic/-mono-iodotyrosine was 4.4.in 1~. HCI whereas I had obtained 8.8. 2. They found that the d/-mono-iodotyrosine crystallized from water with 1 tool of water as revealed by elementary analyses. The writer's analyses agreed with those calculated for a non-hydrated product. 3. In contrast to our positive test they failed to obtain a red color with nitrous acid and ammonia (3). In view of the previous demonstrations (4-6) of the relationship of pepsin tyrosine and protease activity, and of the possibility that our product from pepsin might be an isomer of 3-iodotyrosine, the isolation from an alkaline hydrolysate of mildly iodinated pepsin was performed as previously described (1). d/-mono-iodotyrosine was also synthesized as described by Harington and Pitt Rivers (2). After each had been recrystallized four times their properties were determined and the results compared. The detailed results are reported below but may be summarized here. The specific optical rotation of the freshly isolated product from pepsin was found to be so much less than previously reported that we have some doubts of its being optically active. Nearly all other properties, however, were the same as reported earlier. In addition to the above properties, cross-solubility, cross-partition experiments between immiscible solvents, and the ultraviolet absorption spectra at various pH from which pK were calculated leave little doubt but that the product from pepsin is 3-mono-iodotyrosine. Our synthetic product and that from pepsin both responded positively to the nitrous acid-ammonia color test, but it was possible to. free the synthetic product of its color-giving component. From the analyses of successive extractions with butyl alcohol of an acid hydrolysate of iodinated pepsin, a calculation was made of the quantity 19