Identification of the iodine-sensitive tyrosines in porcine pepsin

Porcine pepsin was iodinated at pH 6.0 and 37° with a 13-fold molar excess of ¹²⁵I]triiodide. Loss of proteolytic activity levelled off at 75 ± 5%, and 2.8 ± 0.1 gram atoms of iodine were incorporated per mole of enzyme. Pepsin, iodinated in this manner, was digested with chymotrypsin. The bulk of the label was located in two peptides with the sequences: Leu-Gly-Gly-Ile-Asp-Ser-Ser-diiodoTyr-Tyr and Ile-Gly-Asp-Glu-Pro-Leu-Asn-iodoTyr. The latter peptide is the N-terminal sequence of pepsin. It is postulated that one or both of these tyrosines may form part of the secondary binding site of pepsin.